Substituted biaryl alkyl amides

ABSTRACT

Disclosed herein are substituted biaryl alkyl amide compounds, methods of synthesizing substituted biaryl alkyl amide compounds and methods of treating diseases and/or conditions with substituted biaryl alkyl amide compounds.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application claims the benefit of priority to U.S. Appl. No.61/548,076, filed Oct. 17, 2011, which is hereby incorporated herein byreference in its entirety.

FIELD

The present application relates to the fields of chemistry, biochemistryand medicine. More particularly, disclosed herein are substituted biarylalkyl amide compounds, pharmaceutical compositions that include one ormore substituted biaryl alkyl amide compounds and methods ofsynthesizing the same. Also disclosed herein are methods of treatingdiseases and/or conditions with substituted biaryl alkyl amidecompounds.

BACKGROUND

The ubiquitin-proteasome system (UPS) plays an important role in manycellular processes. The UPS affects the stability, interactions, andlocalization of many biological proteins. The UPS can be perturbed inmany diseases, such as neoplastic diseases, age-related diseases,neurological diseases, immunological diseases, and infectious diseases.Accordingly, a need exists to develop compounds and compositions thateffectively modulate the UPS.

SUMMARY

Some embodiments disclosed herein relate to a compound of Formula (I) ora pharmaceutically acceptable salt thereof. Some embodiments disclosedherein relate to a pharmaceutical composition that includes one or morecompounds of Formula (I).

Some embodiments disclosed herein relate to a compound of Formula (I)that has the structure of Formula (II), or a pharmaceutically acceptablesalt thereof. Some embodiments disclosed herein relate to a compound ofFormula (I) that has the structure of Formula (III), or apharmaceutically acceptable salt thereof. Some embodiments disclosedherein relate to a compound of Formula (I) that has the structure ofFormula (IV), or a pharmaceutically acceptable salt thereof. Someembodiments disclosed herein relate to a compound of Formula (I) thathas the structure of Formula (V), or a pharmaceutically acceptable saltthereof. Some embodiments disclosed herein relate to a compound ofFormula (I) that has the structure of Formula (VI), or apharmaceutically acceptable salt thereof. Some embodiments disclosedherein relate to a pharmaceutical composition that includes one or morecompounds of Formula (I) that have the structure of Formula (II), (III),(IV), (V), and/or (VI).

Some embodiments disclosed herein relate to a compound of Formula (Ia),(Ib), (Ic) or (Id), or a pharmaceutically acceptable salt thereof, or apharmaceutical composition that includes one or more compounds ofFormula (Ia), (Ib), (Ic) or (Id).

Some embodiments disclosed herein relate to methods for inhibiting theubiquitin-proteasome system in a subject that can include administeringto a subject a compound of Formula (I), (Ia), (Ib), (Ic), (Id), (II),(III), (IV), (V), and/or (VI), or a pharmaceutically acceptable saltthereof, or a pharmaceutical composition that includes one or morecompounds of Formula (I), (Ia), (Ib), (Ic), (Id), (II), (III), (IV),(V), and/or (VI) in a therapeutically effective amount sufficient toinhibit the ubiquitin-proteasome system in the subject. In someembodiments, the subject can be a human.

Some embodiments disclosed herein relate to methods for inhibiting Cdc34in a subject that can include administering to a subject a compound ofFormula (I), (Ia), (Ib), (Ic), (Id), (II), (III), (IV), (V), and/or(VI), or a pharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition that includes one or more compounds of Formula (I), (Ia),(Ib), (Ic), (Id), (II), (III), (IV), (V), and/or (VI) in atherapeutically effective amount sufficient to inhibit Cdc34 in thesubject. In some embodiments, the subject can be a human. In someembodiments, the Cdc34 can be hCdc34.

Some embodiments disclosed herein relate to methods for inhibitingcellular proliferation in a subject comprising administering to thesubject a compound of Formula (I), (Ia), (Ib), (Ic), (Id), (II), (III),(IV), (V), and/or (VI), or a pharmaceutically acceptable salt thereof,or a pharmaceutical composition that includes one or more compounds ofFormula (I), (Ia), (Ib), (Ic), (Id), (II), (III), (IV), (V), and/or (VI)in a therapeutically effective amount sufficient to inhibit cellularproliferation in said subject. In some embodiments, the subject can be ahuman.

Some embodiments disclosed herein relate to methods for ameliorating acondition selected from among a neoplastic disease, a neurologicaldisease, an immunological disease, and an infectious disease that caninclude administering to a subject suffering from the condition atherapeutically effective amount of a compound of Formula (I), (Ia),(Ib), (Ic), (Id), (II), (III), (IV), (V), and/or (VI), or apharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition that includes one or more compounds of Formula (I), (Ia),(Ib), (Ic), (Id), (II), (III), (IV), (V), and/or (VI). In someembodiments, the neoplastic disease can be cancer. In some embodiments,the subject can be a human.

Some embodiments disclosed herein relate to methods for identifying acandidate therapeutic compound that can include determining theeffective amount of a compound of Formula (I), (Ia), (Ib), (Ic), (Id),(II), (III), (IV), (V), and/or (VI), or a pharmaceutically acceptablesalt thereof, on the extent of ubiquitination of p27^(Kip1) by anSCF^(Skp2) E3 complex, such that the compound is identified as acandidate therapeutic compound if the compound significantly reduces theextent of ubiquitination.

Some embodiments disclosed herein relate to methods for determining theeffect of a candidate therapeutic compound that can include determiningthe effective amount of a compound of Formula (I), (Ia), (Ib), (Ic),(Id), (II), (III), (IV), (V), and/or (VI), or a pharmaceuticallyacceptable salt thereof, on the extent of ubiquitin chain initiation orubiquitin chain length, such that the compound is identified as acandidate therapeutic compound if the compound significantly reducessaid extent of ubiquitin chain initiation or ubiquitin chain length.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates the proliferation assay results in the SK-MEL-28(metastatic melanoma) cell line. Compounds B1 through B14 were tested in20 μM and 10 μM concentration respectively.

FIG. 2A illustrates the proliferation assay results in the MDA-MB-468(breast cancer) cell line. Compounds B3, B5, B7, B11 and B12 were testedin 20 μM and 10 μM concentration respectively.

FIG. 2B illustrates the proliferation assay results where Panc-1(pancreatic cancer) cell line was used. Compounds B3, B5, B7, B11 andB12 were tested in 20 μM and 10 μM concentration respectively.

FIG. 3 illustrates TNF-α induced NFkB reporter assay results in theMDA-MB-468-NFkB-Luc2p cell line. Compounds B1 through B14 were tested in20 μM and 5 concentration respectively.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Definitions

As used herein, common organic abbreviations are defined as follows:

Ac Acetyl

-   -   Ac₂O Acetic anhydride    -   aq. Aqueous

Bn Benzyl Bz Benzoyl

-   -   BOC or Boc tert-Butoxycarbonyl    -   Bu n-Butyl    -   cat. Catalytic    -   Cbz Carbobenzyloxy    -   CDI 1,1′-carbonyldiimidazole    -   ° C. Temperature in degrees Centigrade    -   DBU 1,8-Diazabicyclo[5.4.0]undec-7-ene    -   DCE 1,2-Dichloroethane    -   DCM Methylene chloride    -   DIEA Diisopropylethylamine    -   (DHQ)₂PHAL Hydroquinine 1,4-phthalazinediyl diether    -   DMA Dimethylacetamide    -   DME Dimethoxyethane    -   DMF N,N′-Dimethylformamide    -   DMSO Dimethylsulfoxide    -   DPPA Diphenylphosphoryl azide    -   ee % Enantiomeric excess

Et Ethyl

-   -   EtOAc or EA Ethyl acetate    -   g Gram(s)    -   h or hr Hour(s)    -   HATU 2-(1H-7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyl uronium    -   hexafluorophosphate    -   HOBT N-Hydroxybenzotriazole    -   iPr Isopropyl    -   LCMS Liquid chromatography-mass spectrometry    -   LDA Lithium diisopropylamide    -   LiHMDS Lithium bis(trimethylsilyl)amide    -   m or min Minute(s)    -   mCPBA meta-Chloroperoxybenzoic Acid    -   MeOH Methanol    -   MeCN Acetonitrile    -   mL Milliliter(s)    -   MTBE Methyl tertiary-butyl ether    -   NH₄OAc Ammonium acetate    -   NMO N-Methylmorpholine-N-Oxide    -   PE Petroleum ether    -   PG Protecting group    -   Pd/C Palladium on activated carbon    -   Pd(dppf)Cl₂        1,1′-Bis(diphenylphosphino)ferrocene-palladium(II)dichloride    -   Ph Phenyl    -   ppt Precipitate    -   PMBC 4-Methoxybenzyl chloride    -   RCM Ring closing metathesis    -   rt Room temperature    -   sBuLi sec-Butylithium    -   SFC Supercritical fluid chromatography    -   TBAF Tetrabutylammonium fluoride    -   TEA Triethylamine    -   TCDI 1,1′-Thiocarbonyl diimidazole    -   TEMPO 2,2,6,6-Tetramethylpiperidin-1-yl)oxyl    -   Tert, t tertiary    -   TFA Trifluoroacetic acid    -   TFAA Trifluoroacetic acid anhydride    -   THF Tetrahydrofuran    -   TLC Thin-layer chromatography    -   TMEDA Tetramethylethylenediamine    -   TMSNCO trimethylsilyl isocyanate    -   μL Microliter(s)

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of ordinary skillin the art. All patents, applications, published applications and otherpublications referenced herein are incorporated by reference in theirentirety unless stated otherwise. In the event that there are aplurality of definitions for a term herein, those in this sectionprevail unless stated otherwise.

Terms and phrases used in this application, and variations thereof,especially in the appended claims, unless otherwise expressly stated,should be construed as open ended as opposed to limiting. As examples ofthe foregoing, the term ‘including’ should be read to mean ‘including,without limitation,’ ‘including but not limited to,’ or the like; theterm ‘comprising’ as used herein is synonymous with ‘including,’‘containing,’ or ‘characterized by,’ and is inclusive or open-ended anddoes not exclude additional, unrecited elements or method steps; theterm ‘having’ should be interpreted as ‘having at least;’ the term‘includes’ should be interpreted as ‘includes but is not limited to;’the term ‘example’ is used to provide exemplary instances of the item indiscussion, not an exhaustive or limiting list thereof; and use of termslike ‘preferably,’ ‘preferred,’ ‘desired,’ or ‘desirable,’ and words ofsimilar meaning should not be understood as implying that certainfeatures are critical, essential, or even important to the structure orfunction of the invention, but instead as merely intended to highlightalternative or additional features that may or may not be utilized in aparticular embodiment of the invention. In addition, the term“comprising” is to be interpreted synonymously with the phrases “havingat least” or “including at least”. When used in the context of aprocess, the term “comprising” means that the process includes at leastthe recited steps, but may include additional steps. When used in thecontext of a compound or composition, the term “comprising” means thatthe compound or composition includes at least the recited features orcomponents, but may also include additional features or components.Likewise, a group of items linked with the conjunction ‘and’ should notbe read as requiring that each and every one of those items be presentin the grouping, but rather should be read as ‘and/or’ unless expresslystated otherwise. Similarly, a group of items linked with theconjunction ‘or’ should not be read as requiring mutual exclusivityamong that group, but rather should be read as ‘and/or’ unless expresslystated otherwise.

With respect to the use of substantially any plural and/or singularterms herein, those having skill in the art can translate from theplural to the singular and/or from the singular to the plural as isappropriate to the context and/or application. The varioussingular/plural permutations may be expressly set forth herein for sakeof clarity. The indefinite article “a” or “an” does not exclude aplurality. A single processor or other unit may fulfill the functions ofseveral items recited in the claims. The mere fact that certain measuresare recited in mutually different dependent claims does not indicatethat a combination of these measures cannot be used to advantage. Anyreference signs in the claims should not be construed as limiting thescope.

As used herein, any “R” group(s) such as, without limitation, R, R¹, R²,R³, R⁴, R⁵, R⁶, R⁷, R⁸, and R⁹ represent substituents that can beattached to the indicated atom. An R group may be substituted orunsubstituted. If two “R” groups are described as being “taken together”the R groups and the atoms they are attached to can form a cycloalkyl,aryl, heteroaryl or heterocyclyl. For example, without limitation, ifR^(1a) and R^(1b) of an NR^(1a)R^(1b) group are indicated to be “takentogether,” it means that they are covalently bonded to one another toform a ring:

Whenever a group is described as being “optionally substituted” thatgroup may be unsubstituted or substituted with one or more of theindicated substituents. Likewise, when a group is described as being“unsubstituted or substituted” if substituted, the substituent(s) may beselected from one or more the indicated substituents. If no substituentsare indicated, it is meant that the indicated “optionally substituted”or “substituted” group may be substituted with one or more group(s)individually and independently selected from alkyl, alkenyl, alkynyl,cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heteroaryl,heteroalicyclyl, aralkyl, heteroaralkyl, (heteroalicyclyl)alkyl,hydroxy, protected hydroxyl, alkoxy, aryloxy, acyl, mercapto, alkylthio,arylthio, cyano, halogen, thiocarbonyl, O-carbamyl, N-carbamyl,O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, S-sulfonamido,N-sulfonamido, C-carboxy, protected C-carboxy, O-carboxy, isocyanato,thiocyanato, isothiocyanato, nitro, silyl, sulfenyl, sulfinyl, sulfonyl,haloalkyl, haloalkoxy, trihalomethanesulfonyl,trihalomethanesulfonamido, an amino, a mono-substituted amino group anda di-substituted amino group, and protected derivatives thereof.

As used herein, “C_(a) to C_(b)” in which “a” and “b” are integers referto the number of carbon atoms in an alkyl, alkenyl or alkynyl group, orthe number of carbon atoms in the ring of a cycloalkyl, cycloalkenyl,cycloalkynyl or aryl group, or the total number of carbon atoms andheteroatoms in a heteroalkyl, heterocyclyl, heteroaryl orheteroalicyclyl group. That is, the alkyl, alkenyl, alkynyl, ring of thecycloalkyl, ring of the cycloalkenyl, ring of the cycloalkynyl, ring ofthe aryl, ring of the heteroaryl or ring of the heteroalicyclyl cancontain from “a” to “b”, inclusive, carbon atoms. Thus, for example, a“C₁ to C₄ alkyl” group refers to all alkyl groups having from 1 to 4carbons, that is, CH₃—, CH₃CH₂—, CH₃CH₂CH₂—, (CH₃)₂CH—, CH₃CH₂CH₂CH₂—,CH₃CH₂CH(CH₃)— and (CH₃)₃C—. If no “a” and “b” are designated withregard to an alkyl, alkenyl, alkynyl, cycloalkyl cycloalkenyl,cycloalkynyl, aryl, heteroaryl or heteroalicyclyl group, the broadestrange described in these definitions is to be assumed.

As used herein, “alkyl” refers to a straight or branched hydrocarbonchain that comprises a fully saturated (no double or triple bonds)hydrocarbon group. The alkyl group may have 1 to 20 carbon atoms(whenever it appears herein, a numerical range such as “1 to 20” refersto each integer in the given range; e.g., “1 to 20 carbon atoms” meansthat the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3carbon atoms, etc., up to and including 20 carbon atoms, although thepresent definition also covers the occurrence of the term “alkyl” whereno numerical range is designated). The alkyl group may also be a mediumsize alkyl having 1 to 10 carbon atoms. The alkyl group could also be alower alkyl having 1 to 6 carbon atoms. The alkyl group of the compoundsmay be designated as “C₁-C₄ alkyl” or similar designations. By way ofexample only, “C₁-C₄ alkyl” indicates that there are one to four carbonatoms in the alkyl chain, i.e., the alkyl chain is selected from methyl,ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl.Typical alkyl groups include, but are in no way limited to, methyl,ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl andhexyl. The alkyl group may be substituted or unsubstituted.

As used herein, “alkenyl” refers to an alkyl group that contains in thestraight or branched hydrocarbon chain one or more double bonds. Analkenyl group may be unsubstituted or substituted.

As used herein, “alkynyl” refers to an alkyl group that contains in thestraight or branched hydrocarbon chain one or more triple bonds. Analkynyl group may be unsubstituted or substituted.

As used herein, “cycloalkyl” refers to a completely saturated (no doubleor triple bonds) mono- or multi-cyclic hydrocarbon ring system. Whencomposed of two or more rings, the rings may be joined together in afused fashion. Cycloalkyl groups can contain 3 to 10 atoms in thering(s) or 3 to 8 atoms in the ring(s). A cycloalkyl group may beunsubstituted or substituted. Typical cycloalkyl groups include, but arein no way limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl and cyclooctyl.

As used herein, “cycloalkenyl” refers to a mono- or multi- cyclichydrocarbon ring system that contains one or more double bonds in atleast one ring; although, if there is more than one, the double bondscannot form a fully delocalized pi-electron system throughout all therings (otherwise the group would be “aryl,” as defined herein). Whencomposed of two or more rings, the rings may be connected together in afused fashion. A cycloalkenyl group may be unsubstituted or substituted.

As used herein, “cycloalkynyl” refers to a mono- or multi- cyclichydrocarbon ring system that contains one or more triple bonds in atleast one ring. If there is more than one triple bond, the triple bondscannot form a fully delocalized pi-electron system throughout all therings. When composed of two or more rings, the rings may be joinedtogether in a fused fashion. A cycloalkynyl group may be unsubstitutedor substituted.

As used herein, “aryl” refers to a carbocyclic (all carbon) monocyclicor multicyclic aromatic ring system (including fused ring systems wheretwo carbocyclic rings share a chemical bond) that has a fullydelocalized pi-electron system throughout all the rings. The number ofcarbon atoms in an aryl group can vary. For example, the aryl group canbe a C₆-C₁₄ aryl group, a C₆-C₁₀ aryl group, or a C₆ aryl group.Examples of aryl groups include, but are not limited to, benzene,naphthalene and azulene. An aryl group may be substituted orunsubstituted.

As used herein, “heteroaryl” refers to a monocyclic or multicyclicaromatic ring system (a ring system with fully delocalized pi-electronsystem) that contain(s) one or more heteroatoms, that is, an elementother than carbon, including but not limited to, nitrogen, oxygen andsulfur. The number of atoms in the ring(s) of a heteroaryl group canvary. For example, the heteroaryl group can contain 4 to 14 atoms in thering(s), 5 to 10 atoms in the ring(s) or 5 to 6 atoms in the ring(s).Furthermore, the term “heteroaryl” includes fused ring systems where tworings, such as at least one aryl ring and at least one heteroaryl ring,or at least two heteroaryl rings, share at least one chemical bond.Examples of heteroaryl rings include, but are not limited to, furan,furazan, thiophene, benzothiophene, phthalazine, pyrrole, oxazole,benzoxazole, 1,2,3-oxadiazole, 1,2,4-oxadiazole, thiazole,1,2,3-thiadiazole, 1,2,4-thiadiazole, benzothiazole, imidazole,benzimidazole, indole, indazole, pyrazole, benzopyrazole, isoxazole,benzoisoxazole, isothiazole, triazole, benzotriazole, thiadiazole,tetrazole, pyridine, pyridazine, pyrimidine, pyrazine, purine,pteridine, quinoline, isoquinoline, quinazoline, quinoxaline, cinnoline,and triazine. A heteroaryl group may be substituted or unsubstituted.

As used herein, “heterocyclyl” or “heteroalicyclyl” refers to three-,four-, five-, six-, seven-, eight-, nine-, ten-, up to 18-memberedmonocyclic, bicyclic, and tricyclic ring system wherein carbon atomstogether with from 1 to 5 heteroatoms constitute said ring system. Aheterocycle may optionally contain one or more unsaturated bondssituated in such a way, however, that a fully delocalized pi-electronsystem does not occur throughout all the rings. The heteroatom(s) is anelement other than carbon including, but not limited to, oxygen, sulfur,and nitrogen. A heterocycle may further contain one or more carbonyl orthiocarbonyl functionalities, so as to make the definition includeoxo-systems and thio-systems such as lactams, lactones, cyclic imides,cyclic thioimides and cyclic carbamates. When composed of two or morerings, the rings may be joined together in a fused fashion.Additionally, any nitrogens in a heteroalicyclic may be quaternized.Heterocyclyl or heteroalicyclic groups may be unsubstituted orsubstituted. Examples of such “heterocyclyl” or “heteroalicyclyl” groupsinclude but are not limited to, 1,3-dioxin, 1,3-dioxane, 1,4-dioxane,1,2-dioxolane, 1,3-dioxolane, 1,4-dioxolane, 1,3-oxathiane,1,4-oxathiin, 1,3-oxathiolane, 1,3-dithiole, 1,3-dithiolane,1,4-oxathiane, tetrahydro-1,4-thiazine, 2H-1,2-oxazine, maleimide,succinimide, barbituric acid, thiobarbituric acid, dioxopiperazine,hydantoin, dihydrouracil, trioxane, hexahydro-1,3,5-triazine,imidazoline, imidazolidine, isoxazoline, isoxazolidine, oxazoline,oxazolidine, oxazolidinone, thiazoline, thiazolidine, morpholine,oxirane, piperidine N-Oxide, piperidine, piperazine, pyrrolidine,pyrrolidone, pyrrolidione, 4-piperidone, pyrazoline, pyrazolidine,2-oxopyrrolidine, tetrahydropyran, 4H-pyran, tetrahydrothiopyran,thiamorpholine, thiamorpholine sulfoxide, thiamorpholine sulfone, andtheir benzo-fused analogs (e.g., benzimidazolidinone,tetrahydroquinoline, 3,4-methylenedioxyphenyl).

As used herein, “alkoxy” refers to the formula —OR wherein R is analkyl, an alkenyl, an alkynyl, a cycloalkyl, a cycloalkenyl or acycloalkynyl is defined as above. A non-limiting list of alkoxys aremethoxy, ethoxy, n-propoxy, 1-methylethoxy (isopropoxy), n-butoxy,iso-butoxy, sec-butoxy and tert-butoxy. An alkoxy may be substituted orunsubstituted.

As used herein, “acyl” refers to a hydrogen, alkyl, alkenyl, alkynyl, oraryl connected, as substituents, via a carbonyl group. Examples includeformyl, acetyl, propanoyl, benzoyl, and acryl. An acyl may besubstituted or unsubstituted.

As used herein, “hydroxyalkyl” refers to an alkyl group in which one ormore of the hydrogen atoms are replaced by a hydroxy group. Exemplaryhydroxyalkyl groups include but are not limited to, 2-hydroxyethyl,3-hydroxypropyl, 2-hydroxypropyl, and 2,2-dihydroxyethyl. A hydroxyalkylmay be substituted or unsubstituted.

As used herein, “haloalkyl” refers to an alkyl group in which one ormore of the hydrogen atoms are replaced by a halogen (e.g.,mono-haloalkyl, di-haloalkyl and tri-haloalkyl). Such groups include butare not limited to, chloromethyl, fluoromethyl, difluoromethyl,trifluoromethyl and 1-chloro-2-fluoromethyl, 2-fluoroisobutyl. Ahaloalkyl may be substituted or unsubstituted.

As used herein, “aminoalkyl” refers to an optionally substituted aminogroup connected, as a substituent, via a lower alkylene group. Examplesinclude H₂N—O—(CH₂)_(n)—, or (Boc)-NH—(CH₂)_(n)—, wherein n is aninteger in the range of 1 to 6.

As used herein, “haloalkoxy” refers to an alkoxy group in which one ormore of the hydrogen atoms are replaced by a halogen (e.g.,mono-haloalkoxy, di-haloalkoxy and tri- haloalkoxy). Such groups includebut are not limited to, chloromethoxy, fluoromethoxy, difluoromethoxy,trifluoromethoxy and 1-chloro-2-fluoromethoxy, 2-fluoroisobutoxy. Ahaloalkoxy may be substituted or unsubstituted.

As used herein, “aryloxy” and “arylthio” refers to RO— and RS—, in whichR is an aryl, such as but not limited to phenyl. Both an aryloxy andarylthio may be substituted or unsubstituted.

As used herein, “arylalkoxy” refers to an alkoxy group in which one ormore of the hydrogen atoms are replaced by an aryl group (e.g.,—OCH₂Ph).

The term “amino” as used herein refers to a —NH₂ group.

As used herein, the term “hydroxy” refers to a —OH group.

A “cyano” group refers to a “—C≡N” group.

The term “azido” as used herein refers to a —N₃ group.

The term “halogen atom” or “halogen” as used herein, means any one ofthe radio-stable atoms of column 7 of the Periodic Table of theElements, such as, fluorine, chlorine, bromine and iodine.

Where the numbers of substituents is not specified (e.g. haloalkyl),there may be one or more substituents present. For example “haloalkyl”may include one or more of the same or different halogens. As anotherexample, “C₁-C₃ alkoxyphenyl” may include one or more of the same ordifferent alkoxy groups containing one, two or three atoms.

As used herein, the abbreviations for any protective groups, amino acidsand other compounds, are, unless indicated otherwise, in accord withtheir common usage, recognized abbreviations, or the IUPAC-IUBCommission on Biochemical Nomenclature (See, Biochem. 11:942-944 (1972),the disclosure of which is incorporated herein by reference in itsentirety).

The term “pharmaceutically acceptable salt,” especially when referringto a pharmaceutically acceptable salt of the compound of Formula (I),(Ia), (Ib), (Ic), (Id), (II), (III), (IV), (V) or (VI), refers to anypharmaceutically acceptable salts of a compound. Exemplary salts includean acid addition salt of a compound. Pharmaceutical salts can beobtained by reacting a compound with inorganic acids such as hydrohalicacid (e.g., hydrochloric acid or hydrobromic acid), sulfuric acid,nitric acid and phosphoric acid. Pharmaceutical salts can also beobtained by reacting a compound with an organic acid such as aliphaticor aromatic carboxylic or sulfonic acids, for example formic, acetic,succinic, lactic, malic, tartaric, citric, ascorbic, nicotinic,methanesulfonic, ethanesulfonic, p-toluensulfonic, salicylic ornaphthalenesulfonic acid. Pharmaceutical salts can also be obtained byreacting a compound with a base to form a salt such as an ammonium salt,an alkali metal salt, such as a sodium or a potassium salt, an alkalineearth metal salt, such as a calcium or a magnesium salt, a salt oforganic bases such as dicyclohexylamine, N-methyl-D-glucamine,tris(hydroxymethyl)methylamine, C₁-C₇ alkylamine, cyclohexylamine,triethanolamine, ethylenediamine, and salts with amino acids such asarginine and lysine. Exemplary pharmaceutically acceptable salts are thealkali metal salts (sodium or potassium), the alkaline earth metal salts(calcium or magnesium), or ammonium salts derived from ammonia or frompharmaceutically acceptable organic amines, for example C₁-C₇alkylamine, cyclohexylamine, triethanolamine, ethylenediamine ortris-(hydroxymethyl)-aminomethane. With respect to compounds that arebasic amines, exemplary pharmaceutically acceptable salts are acidaddition salts of pharmaceutically acceptable inorganic or organicacids, for example, hydrohalic, sulfuric, phosphoric acid or aliphaticor aromatic carboxylic or sulfonic acid, for example acetic, succinic,lactic, malic, tartaric, citric, ascorbic, nicotinic, methanesulfonic,p-toluensulfonic or naphthalenesulfonic acid.

The term “pharmaceutical composition” refers to a mixture of one or morecompounds disclosed herein with other chemical components, such asdiluents or carriers.

The term “physiologically acceptable” defines a carrier, diluent orexcipient that does not abrogate the biological activity and propertiesof the compound.

As used herein, a “carrier” refers to a compound that facilitates theincorporation of a compound into cells or tissues.

As used herein, an “excipient” refers to an inert substance that isadded to a pharmaceutical composition to provide, without limitation,bulk, consistency, stability, binding ability, lubrication,disintegrating ability etc., to the composition.

A “diluent” is a type of excipient. A “diluent” refers to an ingredientin a pharmaceutical composition that lacks pharmacological activity butmay be pharmaceutically necessary or desirable.

As used herein, the term “subject” refers to a mammal. Exemplary mammalsinclude, but are not limited to, humans, domestic animals (e.g., a dog,cat, or the like), farm animals (e.g., a cow, a sheep, a pig, a horse,or the like) or laboratory animals (e.g., a monkey, a rat, a mouse, arabbit, a guinea pig, or the like).

As used herein, the terms “treating,” “treatment,” “therapeutic,” or“therapy” do not necessarily mean total cure or abolition of the diseaseor condition. Any alleviation of any undesired signs or symptoms of adisease or condition, to any extent can be considered treatment and/ortherapy. Furthermore, treatment may include acts that may worsen thepatient's overall feeling of well-being or appearance.

The term “therapeutically effective amount” is used to indicate anamount of an active compound, or pharmaceutical agent, that elicits abiological or medicinal response.

It is understood that, in any compound described herein having one ormore chiral centers, if an absolute stereochemistry is not expresslyindicated, then each center may independently be of R-configuration orS-configuration or a mixture thereof. Thus, the compounds providedherein may be enantiomerically pure, enantiomerically enriched, racemicmixture, diastereomerically pure, diastereomerically enriched, or astereoisomeric mixture. In addition it is understood that, in anycompound described herein having one or more double bond(s) generatinggeometrical isomers that can be defined as E or Z, each double bond mayindependently be E or Z a mixture thereof. Likewise, it is understoodthat, in any compound described, all tautomeric forms are also intendedto be included.

It is understood that the methods and combinations described hereininclude crystalline forms (also known as polymorphs, which include thedifferent crystal packing arrangements of the same elemental compositionof a compound), amorphous phases, salts, solvates, and hydrates. In someembodiments, the compounds described herein exist in solvated forms withpharmaceutically acceptable solvents such as water, ethanol, or thelike. In other embodiments, the compounds described herein exist inunsolvated form. Solvates contain either stoichiometric ornon-stoichiometric amounts of a solvent, and may be formed during theprocess of crystallization with pharmaceutically acceptable solventssuch as water, ethanol, or the like. Hydrates are formed when thesolvent is water, or alcoholates are formed when the solvent is alcohol.In addition, the compounds provided herein can exist in unsolvated aswell as solvated forms. In general, the solvated forms are consideredequivalent to the unsolvated forms for the purposes of the compounds andmethods provided herein.

Where a range of values is provided, it is understood that the upper andlower limit, and each intervening value between the upper and lowerlimit of the range is encompassed within the embodiments.

II. The Ubiquitin-Proteasome System (UPS)

The ubiquitin-proteasome system (UPS) controls the stability,interactions, and localization of many thousands of proteins acrossvirtually all cellular processes. The UPS degrades damaged proteins andprovides an important biological mechanism for removing dysfunctionalproteins created by transcription, translation and/or folding errors.

The UPS conjugates ubiquitin onto lysine residues of damaged proteins.Ubiquitin is a highly conserved 76-amino acid polypeptide that isabundantly present in eukaryotic cells. Ubiquitin conjugation by the UPSis catalyzed by an enzymatic cascade of at least three classes ofenzymes: (i) E1, (ii) E2, and (iii) E3. The E1 enzyme activatesubiquitin as a thioester and transfers the activated ubiquitin to acatalytic cysteine residue of E2. The E2 enzyme is a serves as aubiquitin carrier. The E3 enzyme is a ubiquitin protein ligase thatattaches the activated ubiquitin to the damaged protein. The resultingubiquitylated protein can be targeted to lysosomes or proteasomes.

In certain diseases the UPS becomes compromised, either leading toexcessive accumulation of unwanted proteins or the abnormal degradationof desired proteins. Perturbation of the UPS is involved in a number ofneoplastic diseases, age-related diseases, neurological diseases,immunological diseases, and infectious diseases. For example, anoveractive UPS can lead to the degradation of desired proteins, such asdegradation of tumor suppressor proteins. Alternatively, an underactiveUPS can lead to the accumulation of undesired proteins, such asaccumulation of oncogenic proteins. A malfunctioning UPS can lead tounregulated cellular proliferation and the formation of one or moreneoplasms. Accordingly, there is a need for compounds and compositionsthat can regulate the UPS. Desired biological properties of suchcompounds and compositions include one or more of the following: highefficacy, high potency, low toxicity, high stability, specificity forUPS components, and a long biological half-life.

III. Compounds

Compounds of Formula (I) can regulate the UPS. For example, compounds ofFormula (I) can inhibit Cdc34, which is the primary E2 enzyme for thecullin-RING ligase (CRL) superfamily class of E3 enzymes. In someembodiments, compounds of Formula (I) can inhibit hCdc34.

Some embodiments disclosed herein relate to a compound of Formula (I) ora pharmaceutically acceptable salt thereof:

-   -   wherein: n can be selected from 0, 1, 2, 3, 4, and 5; each R¹        can be independently selected from halo, cyano, and azido; Z¹,        Z², Z³, and Z⁴ can be each independently —CH— or —N—; R² can be        selected from an optionally substituted (C₁₋₆ alkoxy)C₁₋₆ alkyl,        an optionally substituted C₆₋₁₀ aryl, an optionally substituted        C₅₋₁₀ heteroaryl, an optionally substituted (aryloxy)C₁₋₆ alkyl,        an optionally substituted C₃₋₇ heterocyclyl, an optionally        substituted C₃₋₇ cycloalkyl, an optionally substituted        haloalkyl, and an optionally substituted aminoalkyl; R³ can be        hydrogen or —OH; R⁴ can be selected from an optionally        substituted C₃₋₆ cycloalkyl, an optionally substituted C₃₋₆        heterocyclyl, an optionally substituted C₆₋₁₀ aryl, an        optionally substituted C₅₋₁₀ heteroaryl,

-   -    or R³ and R⁴ together form an optionally substituted C₃₋₆        heterocyclic ring; R⁵ can be hydrogen or —OH; R⁶ can be selected        from —OH, —NHR⁷, an optionally substituted C₁₋₆ alkyl, an        optionally substituted C₁₋₆ alkoxy, an optionally substituted        C₆₋₁₀ aryl, an optionally substituted C₅₋₁₀ heteroaryl, an        optionally substituted aryloxy, or an optionally substituted        arylalkoxy; R⁷ can be hydrogen or an optionally substituted C₁₋₆        alkyl; t can be selected from 0, 1, 2, 3, 4, and 5; provided        that if n is 2, both R¹ are chloro, R² is methoxymethyl, R³ is        —OH, R⁴ is —CH(OH)CH₂OH or

-   -    R⁵ is —OH, and R⁶ is —OH, —NHR⁷, or methoxy; then at least one        of Z¹, Z², Z³, and Z⁴ is —N—; further provided that if n is 2,        both R¹ are chloro, R² is methoxymethyl or phenyl, R³ is        hydrogen, R⁴ is

-   -    R⁵ is hydrogen, and R⁶ is —OH; then at least one of Z¹, Z², Z³,        and Z⁴ is —N—; provided that if n is 2, both R¹ are chloro, R²        is methoxymethyl, and R³ and R⁴ together form an optionally        substituted heterocyclic ring, then at least one of Z¹, Z², Z³,        and Z⁴ is —N—; provided that if n is 2, both R¹ are chloro, R²        is methoxymethyl, R³ is —OH, R⁴ is

-   -    R⁵ is —OH, then R⁶ cannot be —OH, —NH(CH₂)₃OCH₃, or        —NH(CH₂)₃N(CH₃)₂; provided that if n is 2, both R¹ are chloro,        R² is methoxymethyl, R³ is —OH, R⁴ is

-   -    R⁵ is —OH, R⁶ is methoxy, then the compound cannot be

Some embodiments disclosed herein relate to a compound of Formula (I) ora pharmaceutically acceptable salt thereof, wherein: n can be selectedfrom 0, 1, 2, 3, 4, and 5; each R¹ can be independently selected fromhalo, cyano, and azido; Z¹, Z², Z³, and Z⁴ can be each independently—CH— or —N—; R² can be selected from an optionally substituted (C₁₋₆alkoxy)C₁₋₆ alkyl, an optionally substituted C₆₋₁₀ aryl, and anoptionally substituted C₅₋₁₀ heteroaryl; R³ can be hydrogen or —OH; R⁴can be selected from an optionally substituted C₃₋₆ cycloalkyl, anoptionally substituted C₃₋₆ heterocyclyl, an optionally substitutedC₆₋₁₀ aryl, an optionally substituted C₅₋₁₀ heteroaryl, and

R⁵ can be hydrogen or —OH; R⁶ can be selected from —OH, an optionallysubstituted C₁₋₆ alkyl, an optionally substituted C₂₋₆ alkoxy, anoptionally substituted C₆₋₁₀ aryl, an optionally substituted C₅₋₁₀heteroaryl; R⁷ can be hydrogen or an optionally substituted C₁₋₆ alkyl;and provided that if R⁴ is

then R³ and R⁵ cannot be the same.

In some embodiments, n can be selected from the group consisting of 1,2, 3, 4, and 5. In some embodiments, n can be 1 or 2. In someembodiments, n can be 2.

In some embodiments, each R¹ can be independently halo. In someembodiments, each R¹ can be independently chloro, fluoro, or bromo. Insome embodiments, each R¹ can be chloro.

In some embodiments, Z¹ and Z² are each —CH—. In some embodiments, Z³and Z⁴ are each —CH—. In some embodiments, Z³ and Z⁴ are each —N—. Insome embodiments, Z³ can be —CH—, and Z⁴ can be —N—. In someembodiments, Z³ can be —N—, and Z⁴ can be —CH—. In some embodiments,each of Z¹, Z², Z³, and Z⁴ can be —CH—.

In some embodiments, R² can be (C₁₋₆ alkoxy)C₁₋₆ alkyl. In someembodiments, R² can be methoxymethyl. In some embodiments, R² can beethoxymethyl. In some embodiments, R² can be C₆₋₁₀ aryl. In someembodiments, R² can be phenyl. In some embodiments, R² can be C₅₋₁₀heteroaryl. In some embodiments, R² can be furanyl. In some embodiments,R² can be (aryloxy)C₁₋₆ alkyl. In some embodiments, R² can bephenoxymethyl. In some embodiments, R² can be C₃₋₇ heterocyclyl. In someembodiments, R² is selected from optionally substitutedtetrahydrofuranyl or optionally substituted pyrrolidinyl. In someembodiments, the nitrogen atom in pyrrolidinyl is protected with at-butyloxycarbonyl (Boc) protecting group. In some embodiments, R² canbe C₃₋₇ cycloalkyl. In some embodiments, R² can be cyclopentyl. In someembodiments, R² can be haloalkyl. In some embodiments, R² can beselected from —CH₂Cl, —CH₂Br, —CH₂CH₂Cl, —CH₂CH₂Br, —CH(Cl)CH₃ or—CH(Br)CH₃. In some embodiments, R² can be optionally substitutedAminoalkyl. In some embodiments, R² can be selected from —CH₂NH₂,—CH₂NH(Boc), —CH(NH₂)CH₃, and —CH(Boc-NH)CH₃.

In some embodiments, R³ can be hydrogen. In some embodiments, R³ can be—OH.

In some embodiments, R⁴ can be C₃₋₆ cycloalkyl. In some embodiments, R⁴can be cyclohexyl. In some embodiments, R⁴ can be C₃₋₆ heterocyclyl. Insome embodiments, R⁴ can be 1,4,-dioxan-2-one-3-yl. In some embodiments,R⁴ can be C₆₋₁₀ aryl. In some embodiments, R⁴ can be phenyl. In someembodiments, R⁴ can be phenol-2-yl. In some embodiments, R⁴ can be C₅₋₁₀heteroaryl. In some embodiments, R⁴ can be

In some embodiments, R⁴ can be

In some embodiments, R⁴ can be

In some embodiments, R⁵ can be hydrogen. In some embodiments, R⁵ can be—OH.

In some embodiments, R⁶ can be selected from the group consisting of—OH, —NHR⁷, an optionally substituted C₁₋₆ alkoxy, and an optionallysubstituted arylalkoxy. In some embodiments, R⁶ can be —OH. In someembodiments, R⁶ can be —OCH₂Ph. In some embodiments, R⁶ can be —OCH₂CH₃.In some embodiments, R⁶ can be a substituted C₁₋₆ alkyl. In someembodiments, the C₁₋₆ alkyl can be substituted with one or more groupsselected from among halogen, —OH, —COOH, —NR⁸R⁹, C₁₋₆ alkoxy, and C₅₋₁₀heteroaryl; wherein R⁸ and R⁹ are each independently hydrogen or C₁₋₆alkyl. In some embodiments, the C₁₋₆ alkyl can be substituted with —COOHor C₅₋₁₀ heteroaryl. In some embodiments, R⁶ can be C₆₋₁₀ aryl. In someembodiments, R⁶ can be phenyl. In some embodiments, R⁶ can be —NHR⁷.

In some embodiments, R⁷ can be an optionally substituted C₁₋₆ alkyl. Insome embodiments, the C₁₋₆ alkyl can be substituted with one or moregroups selected from C₁₋₆ alkoxy and —NR⁸R⁹; wherein R⁸ and R⁹ are eachindependently hydrogen or C₁₋₆ alkyl.

In some embodiments, R³ and R⁴ together form an optionally substitutedC₃₋₆ heterocyclic ring.

In some embodiments, the compound of Formula (I) can have the structureof Formula (II), or a pharmaceutically acceptable salt thereof:

In some embodiments, the compound of Formula (I) can have the structureof Formula (III), or a pharmaceutically acceptable salt thereof:

In some embodiments, the compound of Formula (I) can have the structureof Formula (IV), or a pharmaceutically acceptable salt thereof:

In some embodiments, the compound of Formula (I) can have the structureof Formula (V), or a pharmaceutically acceptable salt thereof:

In some embodiments, the compound of Formula (I) can have the structureof Formula (VI), or a pharmaceutically acceptable salt thereof:

In some embodiments, R⁴ of Formula (II), Formula (III), Formula (IV),Formula (V) or Formula (VI) can be selected from selected from

Examples of compounds of Formula (I) include, but are not limited to thefollowing:

and pharmaceutically acceptable salts thereof.

In some embodiments, examples of compounds of Formula (I) is selectedfrom Table 1, compounds B1 through B15.

TABLE 1 B1

B2

B3

B4

B5

B6

B7

B8

B9

B10

B11

B12

B13

B14

B15

In some embodiments, examples of compounds of Formula (I) is selectedfrom Table 2, compounds C1 through C5.

TABLE 2 C1

C2

C3

C4

C5

In some embodiments, the exemplary compounds can be enriched withrespect to the shown stereochemistry in an amount>50%, ≧60%, ≧70%, ≧80%,≧90%, ≧95%, or ≧98% as compared to the amount of other stereoisomerimpurities.

Some embodiments disclosed herein relate to a compound of Formula (Ia),(Ib), (Ic) or (Id), or a pharmaceutically acceptable salt thereof:

wherein: n can be selected from 0, 1, 2, 3, 4, and 5; each R¹ can beindependently selected from halo, cyano, and azido; Z¹, Z², Z³, and Z⁴are each independently —CH— or —N—; R² can be selected from anoptionally substituted (C₁₋₆ alkoxy)C₁₋₆ alkyl, an optionallysubstituted C₆₋₁₀ aryl, an optionally substituted C₅₋₁₀ heteroaryl, anoptionally substituted (aryloxy)C₁₋₆ alkyl, an optionally substitutedC₃₋₇ heterocyclyl, an optionally substituted C₃₋₇ cycloalkyl, anoptionally substituted haloalkyl, and an optionally substitutedaminoalkyl; R³ can be hydrogen or —OH; R⁴ can be selected from anoptionally substituted C₃₋₆ cycloalkyl, an optionally substituted C₃₋₆heterocyclyl, an optionally substituted C₆₋₁₀ aryl, an optionallysubstituted C₅₋₁₀ heteroaryl,

or R³ and R⁴ together form an optionally substituted C₃₋₆ heterocyclicring; R⁵ can be hydrogen or —OH; R⁶ can be selected from —OH, —NHR⁷, anoptionally substituted C₁₋₆ alkyl, an optionally substituted C₁₋₆alkoxyl, an optionally substituted C₆₋₁₀ aryl, an optionally substitutedC₅₋₁₀ heteroaryl an optionally substituted aryloxy, or an optionallysubstituted arylalkoxy; R⁷ can be hydrogen or an optionally substitutedC₁₋₆ alkyl; and t can be selected from the group consisting of 0, 1, 2,3, 4, and 5, provided that if n is 2, both R¹ are chloro, R² ismethoxymethyl, R³ is —OH, R⁴ is

R⁵ is —OH, then R⁶ cannot be —OH, —NH(CH₂)₃OCH₃, or —NH(CH₂)₃N(CH₃)₂;provided that if n is 2, both R¹ are chloro, R² is methoxymethyl, R³ is—OH, R⁴ is

R⁵ is —OH, R⁶ is methoxy, then the compound cannot be

Some embodiments disclosed herein relate to a compound of Formula (Ia).Some embodiments disclosed herein relate to a compound of Formula (Ib).Some embodiments disclosed herein relate to a compound of Formula (Ic).Some embodiments disclosed herein relate to a compound of Formula (Id).

Some embodiments disclosed herein relate to a compound of Formula (Ic),or a pharmaceutically acceptable salt thereof, wherein: n can be 2; bothR¹ can be chloro; R² can be methoxymethyl; R³ can be —OH; R⁴ can be

R⁵ can be —OH; R⁶ can be —OH, —NHR⁷, or methoxy; and at least one of Z¹,Z², Z³, and Z⁴ can be —N—.

Some embodiments disclosed herein relate to a compound of Formula (Id),or a pharmaceutically acceptable salt thereof, wherein: n can be 2; bothR¹ can be chloro: R² can be methoxymethyl or phenyl; R³ can be hydrogen;R⁴ can be

R⁵ can be hydrogen; R⁶ can be —OH; and at least one of Z¹, Z², Z³, andZ⁴ can be —N—.

Some embodiments disclosed herein relate to a compound of Formula (Ia)or (Ic), or a pharmaceutically acceptable salt thereof, wherein: n canbe 2; both R¹ are chloro; R² can be methoxymethyl; R³ and R⁴ togetherform an optionally substituted heterocyclic ring; and at least one ofZ¹, Z², Z³, and Z⁴ can be —N—.

In some embodiments, n can be selected from the group consisting of 1,2, 3, 4, and 5. In some embodiments, n can be 1 or 2. In someembodiments, n can be 2.

In some embodiments, each R¹ can be independently halo. In someembodiments, each R¹ can be independently chloro, fluoro, or bromo. Insome embodiments, each R¹ can be chloro.

In some embodiments, Z¹ and Z² are each —CH—. In some embodiments, Z³and Z⁴ are each —CH—. In some embodiments, Z³ and Z⁴ are each —N—. Insome embodiments, Z³ can be —CH—, and Z⁴ can be —N—. In someembodiments, Z³ can be —N—, and Z⁴ can be —CH—. In some embodiments,each of Z¹, Z², Z³, and Z⁴ can be —CH—.

In some embodiments, R² can be (C₁₋₆ alkoxy)C₁₋₆ alkyl. In someembodiments, R² can be methoxymethyl. In some embodiments, R² can beethoxymethyl. In some embodiments, R² can be C₆₋₁₀ aryl. In someembodiments, R² can be phenyl. In some embodiments, R² can be C₅₋₁₀heteroaryl. In some embodiments, R² can be furanyl. In some embodiments,R² can be (aryloxy)C₁₋₆ alkyl. In some embodiments, R² can bephenoxymethyl. In some embodiments, R² can be C₃₋₇ heterocyclyl. In someembodiments, R² is selected from optionally substitutedtetrahydrofuranyl or optionally substituted pyrrolidinyl. In someembodiments, the nitrogen atom in pyrrolidinyl is protected with at-butyloxycarbonyl (Boc) protecting group. In some embodiments, R² canbe C₃₋₇ cycloalkyl. In some embodiments, R² can be cyclopentyl. In someembodiments, R² can be haloalkyl. In some embodiments, R² can beselected from —CH₂Cl, —CH₂Br, —CH₂CH₂Cl, —CH₂CH₂Br, —CH(Cl)CH₃ or—CH(Br)CH₃.

In some embodiments, R² can be optionally substituted aminoalkyl. Insome embodiments, R² can be selected from —CH₂NH₂, —CH₂NH(Boc),—CH(NH₂)CH₃, and —CH(Boc-NH)CH₃.

In some embodiments, R³ can be hydrogen. In some embodiments, R³ can be—OH.

In some embodiments, R⁴ can be C₃₋₆ cycloalkyl. In some embodiments, R⁴can be cyclohexyl. In some embodiments, R⁴ can be C₃₋₆ heterocyclyl. Insome embodiments, R⁴ can be 1,4,-dioxan-2-one-3-yl. In some embodiments,R⁴ can be C₆₋₁₀ aryl. In some embodiments, R⁴ can be phenyl. In someembodiments, R⁴ can be phenol-2-yl. In some embodiments, R⁴ can be C₅₋₁₀heteroaryl. In some embodiments, R⁴ can be

In some embodiments, R⁴ can be

In some embodiments, R⁴ can be

In some embodiments, R⁵ can be hydrogen. In some embodiments, R⁵ can be—OH.

In some embodiments, R⁶ can be selected from the group consisting of—OH, —NHR⁷, an optionally substituted C₁₋₆ alkoxy, and an optionallysubstituted arylalkoxy. In some embodiments, R⁶ can be —OH. In someembodiments, R⁶ can be —OCH₂Ph. In some embodiments, R⁶ can be —OCH₂CH₃.In some embodiments, R⁶ can be a substituted C₁₋₆ alkyl. In someembodiments, the C₁₋₆ alkyl can be substituted with one or more groupsselected from among halogen, —OH, —COOH, —NR⁸R⁹, alkoxy, and C₅₋₁₀heteroaryl; wherein R⁸ and R⁹ are each independently hydrogen or C₁₋₆alkyl. In some embodiments, the C₁₋₆ alkyl can be substituted with —COOHor C₅₋₁₀ heteroaryl. In some embodiments, R⁶ can be C₆₋₁₀ aryl. In someembodiments, R⁶ can be phenyl. In some embodiments, R⁶ can be —NHR⁷.

In some embodiments, R⁷ can be an optionally substituted C₁₋₆ alkyl. Insome embodiments, the C₁₋₆ alkyl can be substituted with one or moregroups selected from alkoxy and —NR⁸R⁹; wherein R⁸ and R⁹ are eachindependently hydrogen or C₁₋₆ alkyl.

In some embodiments, R³ and R⁴ together form an optionally substitutedC₃₋₆ heterocyclic ring.

In some embodiments, the compound of Formula (I)-(VI) (which can includea compound of Formula (Ia), (Ib), (Ic), or (Id)) can be enriched in the(R) or (S) enantiomer or diastereomer with respect to any chiral carbonatom. For example, a compound of Formula (I)-(VI) can be enriched in the(R) or (S) configuration at any chiral carbon atom in an amount>50%,≧60%, ≧70%, ≧80%, ≧90%, ≧95%, or ≧98% compared to the amount of theother of the (R) or (S) configuration. Some compounds of Formula(I)-(VI) can be enriched in a diastereomer having two or more chiralcarbon atoms. In some embodiments, the compound can include a singleenantiomer or diastereomer of a compound of Formula (I) at aconcentration of greater than 99% compared to the total concentration ofthe other enantiomers or diastereomers. In other embodiments, thecompound can include a mixture of two or more diastereomers. Forexample, the compound can include a concentration of one diastereomerof >50%, ≧60%, ≧70%, ≧80%, ≧90%, ≧95%, or ≧98%, as compared to the totalconcentration of the other diastereomers. In some embodiments, thecompound includes a 1:1 mixture of two diastereomers.

With respect to compounds of Formula (I) and (II), in some embodiments,the compounds are enriched in the (R) or (S) enantiomer or diastereomerwith respect to any chiral carbon. In some embodiments, the compound ofFormula (I) or (II) can be enriched in the (R) or (S) configuration atthe sp³ hybridized carbon connected to the nitrogen atom of the—NHC(═O)— group. In some embodiments, the enrichment can be >50%, ≧60%,≧70%, ≧80%, ≧90%, ≧95%, or ≧98%, as compared to the total concentrationof other stereoisomers.

In some embodiments, the compound of Formula (I) or (II) can be enrichedin the (R) or (S) configuration at the carbon connected to R³ and R⁴. Insome embodiments, the enrichment can be >50%, ≧60%, ≧70%, ≧80%, ≧90%,≧95%, or ≧98%, as compared to the total concentration of otherstereoisomers.

In some embodiments, when R⁴ of Formula (I) or (II) is

then the compound of Formula (I) or (II) can be enriched in the (R) or(S) configuration at the carbon connected to R⁵ and (CH₂)_(t)—OH or thecarbon connected to R⁵ and C(═O)R⁶. In some embodiments, the enrichmentcan be >50%, ≧60%, ≧70%, ≧80%, ≧90%, ≧95%, or ≧98%, as compared to thetotal concentration of other stereoisomers.

In some embodiments, the compound of Formula (I) or (II) can be enrichedin the (R) or (S) configuration at the sp³ hybridized carbon connectedto the nitrogen atom of the —NHC(═O)— group; and the compound can beenriched in the (R) or (S) configuration at the carbon connected to R³and R⁴. In some embodiments, the enrichment can be >50%, ≧60%, ≧70%,≧80%, ≧90%, ≧95%, or ≧98%, as compared to the total concentration ofother stereoisomers.

In some embodiments, when R⁴ of Formula (I) or (II) is

then the compound of Formula (I) or (II) can be enriched in the (R) or(S) configuration at the carbon connected to R⁵ and (CH₂)_(t)—OH or thecarbon connected to R⁵ and C(═O)R⁶; and the compound can be enriched inthe (R) or (S) configuration at the sp³ hybridized carbon connected tothe nitrogen atom of the —NHC(═O)-group. In some embodiments, theenrichment can be >50%, ≧60%, ≧70%, ≧80%, ≧90%, ≧95%, or ≧98%, ascompared to the total concentration of other stereoisomers.

In some embodiments, when R⁴ of Formula (I) or (II) is

then the compound of Formula (I) or (II) can be enriched in the (R) or(S) configuration at the carbon connected to R⁵ and (CH₂)_(t)—OH or thecarbon connected to R⁵ and C(═O)R⁶; and the compound can be enriched inthe (R) or (S) configuration at the carbon connected to R³ and R⁴. Insome embodiments, the enrichment can be >50%, ≧60%, ≧70%, ≧80%, ≧90%,≧95%, or ≧98%, as compared to the total concentration of otherstereoisomers.

In some embodiments, when R⁴ of Formula (I) or (II) is

then the compound of Formula (I) or (II) can be enriched in the (R) or(S) configuration at the carbon connected to R⁵ and (CH₂)_(t)—OH or thecarbon connected to R⁵ and C(═O)R⁶; and the compound can be enriched inthe (R) or (S) configuration at the sp³ hybridized carbon connected tothe nitrogen atom of the —NHC(═O)-group; and the compound can beenriched in the (R) or (S) configuration at the carbon connected to R³and R⁴. In some embodiments, the enrichment can be >50%, ≧60%, ≧70%,≧80%, ≧90%, ≧95%, or ≧98%, as compared to the total concentration ofother stereoisomers.

In some embodiments, the compound of Formula (III)-(VI), (Ia), (Ib),(Ic), or (Id) can be enriched with respect to the shown stereochemistryin an amount>50%, ≧60%, ≧70%, ≧80%, ≧90%, ≧95%, or ≧98% as compared tothe amount of other stereoisomer impurities. In some embodiments, thecompound can be enriched with respect to the shown stereochemistry in anamount>99% as compared to the amount of other stereoisomer impurities.

IV. Screening

Compounds of Formula (I)-(VI) (which can include a compound of Formula(Ia), (Ib), (Ic) or (Id)) can be screened for their ability to inhibitCdc34. In some embodiments, the compounds of Formula (I)-(IV) can bescreened by using a high-throughput (HTP) compatible assay. This assaycan be based on ubiquitination of the human cyclin-dependent kinase(CDK) inhibitor p27^(Kip1) by SCF^(Skp2), which is a Skp1-Cullin1-F-box(SCF) E3 complex. The SKP2 locus is often amplified and overexpressed inhuman cancer, thus SCF^(Skp2) is a candidate for therapeuticintervention.

The HTP assay can contain biotinylated-ubiquitin, the E1 enzyme Uba1,the E2 enzyme hCcd34, the SCF^(Skp2) complex, cyclin-dependent kinaseregulatory subunit 1 (Cks1), and p27^(Kip1) that is phosphorylated bycyclin E-Cdk2. Ubiquitination of p27^(Kip1) can be assessed by captureonto an anti- p27^(Kip1) antibody affinity surface and quantitativedetection with a europium-streptavidin conjugate. See Ceccarelli et al.,“An allosteric inhibitor of the human Cdc34 ubiquitin-conjugatingenzyme,” Cell 145:1075-1087, 2011, the disclosure of which isincorporated herein by reference in its entirety.

If the compound of Formula (I)-(VI) significantly reduces the extent ofubiquitination of p27^(Kip1), then the compound can be a candidatetherapeutic compound. If the compound of Formula (I)-(VI) significantlyreduces the extent of ubiquitin chain initiation or ubiquitin chainlength on p27^(Kip1), then the compound can be a candidate therapeuticcompound. Methods for screening compounds of Formula (I)-(VI) for theireffects on the UPS are described in Ceccarelli et al., “An allostericinhibitor of the human Cdc34 ubiquitin-conjugating enzyme,” Cell145:1075-1087, 2011, the disclosure of which is incorporated herein byreference in its entirety.

V. Properties

For more than a decade, researchers have been developing ways to targetthe ubiquitin-proteasome pathway in cancer. However, only one marketedtherapy actually acts on this pathway, which is the proteasome inhibitorVelcade bortezomib([(1R)-3-methyl-1-({(2S)-3-phenyl-2-[(pyrazin-2-ylcarbonyl)amino]propanoyl}-amino)butyl]boronicacid) that is marketed by Takeda Pharmaceutical Co. Ltd.'s MillenniumPharmaceuticals to treat multiple myeloma and mantle cell lymphoma.Kotz, “Celgene skips SKP2,” SciBX 4(28) 2011, the disclosure of which isincorporated herein by reference in its entirety. Velcade's broadside-effect profile has prompted a search for better alternatives.

Researchers are interested in targeting the ubiquitin-proteasome pathwayat points upstream from the proteasome. Ceccarelli et al. identified asmall number of compounds that are able to somewhat regulate theubiquitin-proteasome system (UPS). Ceccarelli et al., “An allostericinhibitor of the human Cdc34 ubiquitin-conjugating enzyme,” Cell145:1075-1087, 2011. Specifically, Ceccarelli et al. demonstrated thatcompound CC0651 inhibits the E2 ubiquitin-conjugating enzyme hCdc34.However, the Ceccarelli team thought that it was too difficult tooptimize CC0651 for the clinic and subsequently abandoned their program.Kotz, “Celgene skips SKP2,” SciBX 4(28) 2011.

CC0651 is only a first step toward targeting an E2 enzyme, and a muchmore potent drug-like molecule is needed for clinical therapy. Kotz,“Celgene skips SKP2,” SciBX 4(28) 2011 at page 1. Furthermore, DavidWebb, the Vice President of research at Celgene stated that the companydoes not plan to pursue CC0651 as a drug lead or Cdc34 target because“it was difficult to see a way forward to get below a micromolar IC₅₀.”Id. Webb goes on to state that “The chemistry didn't look to us like itwas going to get us to the potency needed for a drug.” Id. Celgene hadworked on the project for seven years, used an 11-protein assay,screened an enormous library twice, and could not advance any of theresulting compounds. Id. After two failed attempts to produce ananomolar-potency inhibitor, the Celgene program was stopped. Id. “Theubiquitin ligases themselves are incredibly difficult to drug; weconsider them relatively undruggable,” said Webb. Id.

VI. Synthesis

The compounds described herein can be prepared in various ways. Generalsynthetic routes to the compound of Formulae (I)-(VI) (which can includea compound of Formula (Ia), (Ib), (Ic) or (Id)), and some examples ofmaterials and intermediates used to synthesize the compounds of Formulae(I)-(VI) are shown in Schemes 1-10, and described herein. The routesshown and described herein are illustrative only and are not intended,nor are they to be construed, to limit the scope of the claims in anymanner whatsoever. Those skilled in the art will be able to recognizemodifications of the disclosed syntheses and to devise alternate routesbased on the disclosures herein; all such modifications and alternateroutes are within the scope of the claims.

Some methods for preparing a compound of Formula (E) are shown inScheme 1. In Scheme 1, Z¹, Z², Z³, Z⁴, R¹ and n can be the same as Z¹,Z², Z³, Z⁴, R¹ and n as described herein for Formula (I). The compoundof Formula (A) can be a racemic mixture of enantiomers. In someembodiments, the compound of Formula (A) can be stereospecific, havingan (R) configuration about carbon ‘a’. In some embodiments, the compoundof Formula (A) can be stereospecific, having an (S) configuration aboutcarbon ‘a’. A compound of Formula (B^(I)) can be prepared by reacting acompound of Formula (A) with HPO(OEt)₂. A compound of Formula (B^(II))can be prepared by reacting a compound of Formula (A) with methanesulfonyl chloride (MsCl).

A compound of Formula (D) can be prepared by reacting a compound ofFormula (B^(I)), (B^(II)), or (B^(III)) with a compound of Formula (C).The reaction of a compound of Formula (B^(I)), (B^(II)), or (B^(III))with a compound of Formula (C) results in inversion of thestereochemistry about carbon ‘a’. A compound of Formula (E) can beprepared by reducing a compound of Formula (D). Suitable reducing agentsinclude, but are not limited to, diisobutylaluminium hydride (DIBAL),lithium aluminium hydride (LiAlH₄), and sodium borohydride (NaBH₄).

Some methods for preparing a compound of Formula (L) or (M) are shown inScheme 2. In Scheme 2, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵, and n can be thesame as Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ and n as described herein forFormula (I). In Scheme 2, R⁶ can be an optionally substituted C₁₋₆alkyl, an optionally substituted C₁₋₆ alkoxyl, an optionally substitutedC₆₋₁₀ aryl or an optionally substituted C₅₋₁₀ heteroaryl. In Scheme 2,the compounds can be a racemic mixture of enantiomers or diastereomers.In some embodiments, the compounds can be stereospecific, with carbons‘a’, ‘b’, and ‘c’ each independently having either an (R) or (S)configuration. The compound of Formula (L) can be a compound of

Formula (I), where R⁴ of Formula (I) is

A compound of Formula (G) can be prepared by reacting a compound ofFormula (E) with a compound of Formula (F). A compound of Formula (H)can be prepared by removing the t-butyloxycarbonyl (Boc) protectinggroup from a compound of Formula (G). Suitable deprotection reagentsinclude, but are not limited to, trifluoroacetic acid (TFA),hydrochloric acid, and sulfuric acid. A compound of Formula (K) can beprepared by reacting a compound of Formula (H) with a compound ofFormula (J).

A compound of Formula (L) can be prepared by reducing a compound ofFormula (K), with H₂ such that R³ and R⁵ are both hydrogen.

Alternatively, a compound of Formula (L) can be prepared by reacting acompound of Formula (K) with OsO₄ or KMnO₄, such that R³ and R⁵ are bothhydroxyl. In some embodiments, reacting a compound of Formula (K) withOsO₄ or KMnO₄ can result in isolating a compound of Formula (L) havingan (R,R) configuration about carbons ‘b’ and ‘c’, respectively. In someembodiments, reacting a compound of Formula (K) with OsO₄ or KMnO₄ canresult in isolating a compound of Formula (L) having an (S,S)configuration about carbons ‘b’ and ‘c’, respectively. For example,reacting a compound of Formula (K) with OsO₄ in the presence ofhydroquinine 1,4-phthalazinediyl diether ((DHQ)₂PHAL), K₃Fe(CN)₆, andMeSO₂NH₂ can result in syn addition about the carbon-carbon double bond.

In some embodiments, reacting a compound of Formula (K) with OsO₄ canresult in isolating a compound of Formula (L) having an (R,S)configuration about carbons ‘b’ and ‘c’, respectively. In someembodiments, reacting a compound of Formula (K) with OsO₄ can result inisolating a compound of Formula (L) having an (S,R) configuration aboutcarbons ‘b’ and ‘c’, respectively. For example, reacting a compound ofFormula (K) with OsO₄ in the presence of N-methylmorpholine-N-oxide(NMO) followed by treatment with NaHSO₃ can result in anti additionabout the carbon-carbon double bond.

A compound of Formula (M) can be prepared by hydrolyzing a compound ofFormula (L). Suitable hydrolyzing reagents include, but are not limitedto, NaOH, KOH, LiOH, KHCO₃, H₂SO₄, and HCl.

Some methods for preparing a compound of Formula (M) are shown in Scheme3. In Scheme 3, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ and n can be the same asZ¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ and n as described herein for Formula(I). In Scheme 3, R⁶ can be an optionally substituted C₁₋₆ alkyl, anoptionally substituted C₁₋₆ alkoxyl, an optionally substituted C₆₋₁₀aryl or an optionally substituted C₅₋₁₀ heteroaryl. In Scheme 3, thecompounds can be a racemic mixture of enantiomers or diastereomers. Insome embodiments, the compounds can be stereospecific, with carbons ‘a’,‘b’, and ‘c’ each independently having either an (R) or (S)configuration. The compound of Formula (M) can be a compound of Formula(I), where R⁴ of Formula (I) is

and R⁶ is —OH.

A compound of Formula (N) can be prepared by reacting a compound ofFormula (B^(IV)) with a compound of Formula (J). A compound of Formula(O) can be prepared by reacting a compound of Formula (N) with acompound of Formula (C). The reaction of a compound of Formula (N) witha compound of Formula (C) results in inversion of the stereochemistryabout carbon ‘a’.

A compound of Formula (P) can be prepared by reducing a compound ofFormula (O). Suitable reducing agents include, but are not limited to,diisobutylaluminium hydride (DIBAL), lithium aluminium hydride (LiAlH₄),and sodium borohydride (NaBH₄). A compound of Formula (K) can beprepared by reacting a compound of Formula (P) with a compound ofFormula (F). A compound of Formula (Q) can be prepared by epoxidation ofa compound of Formula (K). Suitable epoxidation reagents include, butare not limited to, a peroxyacid (e.g. meta-chloroperoxybenzoic acid(MCPBA), hydrogen peroxide, and alkyl hydroperoxides (e.g. t-butylhydroperoxide, and ethylbenzene hydroperoxide).

A compound of Formula (M) can be prepared by reacting of a compound ofFormula (Q) with aqueous acid, such that R³ and R⁵ are both hydroxy. Insome embodiments, reacting a compound of Formula (Q) with aqueous acidcan result in isolating a compound of Formula (M) having an (R,S)configuration about carbons ‘b’ and ‘c’, respectively. In someembodiments, reacting a compound of Formula (Q) with aqueous acid canresult in isolating a compound of Formula (M) having an (S,R)configuration about carbons ‘b’ and ‘c’, respectively.

Alternatively, a compound of Formula (M) can be prepared by reacting acompound of Formula (Q) with SmI₂ and dimethylethanolamine (DMAE),followed by NaOH, such that R³ is hydroxyl and R⁵ is hydrogen. In someembodiments, reacting a compound of Formula (Q) with SmI₂ anddimethylethanolamine (DMAE), followed by NaOH can result in isolating acompound of Formula (M) having an (R) configuration about carbon ‘b’. Insome embodiments, reacting a compound of Formula (Q) with SmI₂ anddimethylethanolamine (DMAE), followed by NaOH can result in isolating acompound of Formula (M) having an (S) configuration about carbon ‘b’.

One method for preparing a compound of Formula (L) or (M) is shown inScheme 4. In Scheme 4, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ and n can be thesame as Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ and n as described herein forFormula (I). In Scheme 4, R⁶ can be an optionally substituted C₁₋₆alkyl, an optionally substituted C₁₋₆ alkoxyl, an optionally substitutedC₆₋₁₀ aryl or an optionally substituted C₅₋₁₀ heteroaryl. In Scheme 4,the compounds can be a racemic mixture of enantiomers or diastereomers.In some embodiments, the compounds can be stereospecific, with carbons‘a’ and ‘c’ each independently having either an (R) or (S)configuration. The compound of Formula (L) can be a compound of Formula(I), where R⁴ of Formula (I) is

A compound of Formula (L) can be prepared by reacting of a compound ofFormula (K) with BH₃.SMe₂ followed by aqueous NaOH, such that R³ ishydrogen and R⁵ is hydroxy. In some embodiments, reacting a compound ofFormula (L) with BH₃.SMe₂ followed by aqueous NaOH can result inisolating a compound of Formula (L) having an (R) configuration aboutcarbon ‘c’. In some embodiments, reacting a compound of Formula (L) withBH₃.SMe₂ followed by aqueous NaOH can result in isolating a compound ofFormula (L) having an (S) configuration about carbon ‘c’.

A compound of Formula (M) can be prepared by hydrolyzing a compound ofFormula (L). Suitable hydrolyzing reagents include, but are not limitedto, NaOH, KOH, LiOH, KHCO₃, H₂SO₄, and HCl.

A method for preparing a compound of Formula (M′) is shown in Scheme 5.In Scheme 5, Z¹, Z², Z³, Z⁴, R¹ and n can be the same as Z¹, Z², Z³, Z⁴,R¹ and n as described herein for Formula (I). In Scheme 5, the compoundscan be a racemic mixture of enantiomers or diastereomers. In someembodiments, the compounds can be stereospecific, with carbon ‘a’ havingeither an (R) or (S) configuration, and carbons ‘b’ and ‘c’ having theshown stereochemistry. The compound of Formula (M′) can be a compound ofFormula (I), where R⁴ of Formula (I) is

and R³, R⁵, and R⁶ are each hydroxyl.

A compound of Formula (R) can be prepared by reacting a compound ofFormula (B^(V)) with a compound of Formula (C). The reaction of acompound of Formula (B^(V)) with a compound of Formula (C) results ininversion of the stereochemistry about carbon ‘a’. A compound of Formula(S) can be prepared by reducing a compound of Formula (R). Suitablereducing agents include, but are not limited to, diisobutylaluminiumhydride (DIBAL), lithium aluminium hydride (LiAlH₄), and sodiumborohydride (NaBH₄).

A compound of Formula (T) can be prepared by reacting a compound ofFormula (S) with 1-hydroxypropan-2-one. A compound of Formula (U) can beprepared by reacting a compound of Formula (T) with Br₂ in aqueous NaOH,followed by treatment with benzyl bromide. A compound of Formula (V) canbe prepared by reacting a compound of Formula (U) with2,2-dimethoxypropane (DMP) in acetone.

A compound of Formula (W) can be prepared by removing thebenzyloxycarbonyl (Cbz) protecting group from a compound of Formula (V).Suitable deprotection reagents include, but are not limited to,catalytic hydrogenolysis, KOH, and HBr. A compound of Formula (X) can beprepared by reacting a compound of Formula (W) with a compound ofFormula (J). A compound of Formula (M′) can be prepared by reacting of acompound of Formula (X) with aqueous acid.

Some methods for preparing a compound of Formula (AF) are shown inScheme 6. In Scheme 6, Z¹, Z², Z³, Z⁴, R¹ and n can be the same as Z¹,Z², Z³, Z⁴, R¹ and n as described herein for Formula (I). In Scheme 6,the compounds can be a racemic mixture of enantiomers or diastereomers.In some embodiments, the compounds can be stereospecific, with carbons‘a’ and ‘b’ each independently having either an (R) or (S)configuration.

A compound of Formula (Z) can be prepared by reacting a compound ofFormula (Y) with CuSO₄ in acetone, followed by treatment with K₂CO₃ andH₂O₂, and followed by treatment with ethyl iodide. A compound of Formula(AA) can be prepared by reacting a compound of Formula (Z) with2-chloroacetic acid, triphenylphosphine, and diisopropylazodicarboxylate (DIAD), followed by treatment with triethylamine inethanol. A compound of Formula (AB) can be prepared by reacting acompound of Formula (AA) with benzyl bromide and silver oxide. Acompound of Formula (AD) having (S) stereochemistry at carbon ‘b’ can beprepared by reducing a compound of Formula (AB). Suitable reducingagents include, but are not limited to, diisobutylaluminium hydride(DIBAL), lithium aluminium hydride (LiAlH₄), and sodium borohydride(NaBH₄).

A compound of Formula (AC) can be prepared by reacting a compound ofFormula (Z) with benzyl bromide and silver oxide. A compound of Formula(AD) having (R) stereochemistry at carbon ‘b’ can be prepared byreducing a compound of Formula (AC). Suitable reducing agents include,but are not limited to, diisobutylaluminium hydride (DIBAL), lithiumaluminium hydride (LiAlH₄), and sodium borohydride (NaBH₄).

A compound of Formula (AF) can be prepared by reacting a compound ofFormula (AD) with a compound of Formula (AE).

Methods for preparing a compound of Formula (AM) or (M″) are shown inScheme 7. In Scheme 7, Z¹, Z², Z³, Z⁴, R¹, R² and n can be the same asZ¹, Z², Z³, Z⁴, R¹, R² and n as described herein for Formula (I). InScheme 7, the compounds can be a racemic mixture of enantiomers ordiastereomers. In some embodiments, the compounds can be stereospecific,with carbons ‘a’ and ‘b’ each independently having either an (R) or (S)configuration. The compound of Formula (AM) can be a compound of Formula(I), where R⁴ of Formula (I) is

R³ and R⁵ are each hydroxyl, and R⁶ is methoxy. The compound of Formula(M″) can be a compound of Formula (I), where R⁴ of Formula (I) is

and R³, R⁵ and R⁶ are each hydroxy.

A compound of Formula (AG) can be prepared by reacting a compound ofFormula (AF) with methanesulfonyl chloride (MsCl). A compound of Formula(AH) can be prepared by reacting a compound of Formula (AG) with sodiumazide. A compound of Formula (AI) can be prepared by reducing a compoundof Formula (AH) with a reducing agent such as LiBH₄. A compound ofFormula (AJ) can be prepared by reacting a compound of Formula (AI) witha compound of Formula (J).

A compound of Formula (AK) can be prepared by removing the acetonideprotecting group from a compound of Formula (AJ). Suitable deprotectionreagents include, but are not limited to, H₂SO₄, HCl, acetic acid,trifluoroacetic acid, and cation exchange resins. A compound of Formula(AL) can be prepared by reacting a compound of Formula (AK) with2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO), N-chlorosuccinimide (NCS),and tetrabuylammonium chloride (TBACl), followed by treatment withNaClO₂, NaH₂PO₄, and H₂O₂, followed by treatment with CH₂N₂.

A compound of Formula (AM) can be prepared by removing the benzylprotecting group from a compound of Formula (AL). Suitable deprotectionconditions include, for example, catalytic hydrogenolysis. A compound ofFormula (M″) can be prepared by hydrolyzing a compound of Formula (AM).Suitable hydrolyzing reagents include, but are not limited to, NaOH,KOH, LiOH, KHCO₃, H₂SO₄, and HCl.

Methods for preparing a compound of Formula (AQ) are shown in Scheme 8.In Scheme 8, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵ n and t can be the same asZ¹, Z², Z³, Z⁴, R¹, R², R³, R⁵, n and t as described herein for Formula(I). In Scheme 8, the compounds can be a racemic mixture of enantiomersor diastereomers. In some embodiments, the compounds can bestereospecific, with carbons ‘a’, ‘b’ and ‘c’ each independently havingeither an (R) or (S) configuration. The compound of Formula (AQ) can bea compound of Formula (I), where R⁴ of Formula (I) is

A compound of Formula (AO) can be prepared by reacting a compound ofFormula (E) with a compound of Formula (AN). A compound of Formula (AP)can be prepared by removing the t-butyloxycarbonyl (Boc) protectinggroup from a compound of Formula (AO). Suitable deprotection reagentsinclude, but are not limited to, trifluoroacetic acid (TFA),hydrochloric acid, and sulfuric acid. A compound of Formula (AQ) can beprepared by reacting a compound of Formula (AP) with a compound ofFormula (J).

Methods for preparing a compound of Formula (AW) are shown in Scheme 9.In Scheme 9, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁴ and n can be the same as Z¹,Z², Z³, Z⁴, R¹, R², R³, R⁴ and n as described herein for Formula (I). InScheme 9, the compounds can be a racemic mixture of enantiomers ordiastereomers. In some embodiments, the compounds can be stereospecific,with carbons ‘a’ and ‘b’ each independently having either an (R) or (S)configuration. The compound of Formula (AW) can be a compound of Formula(I), where R⁴ of Formula (I) is an optionally substituted C₃₋₆cycloalkyl, an optionally substituted C₃₋₆ heterocyclyl, an optionallysubstituted C₆₋₁₀ aryl, or an optionally substituted C₅₋₁₀ heteroaryl.

A compound of Formula (AS) where R³ is hydroxyl can be prepared byreacting a compound of Formula (E) with a compound of Formula (AR). Acompound of Formula (AT) where R³ is hydroxyl can be prepared byremoving the t-butyloxycarbonyl (Boc) protecting group from a compoundof Formula (AS). Suitable deprotection reagents include, but are notlimited to, trifluoroacetic acid (TFA), hydrochloric acid, and sulfuricacid. A compound of Formula (AW) where R³ is hydroxyl can be prepared byreacting a compound of Formula (AT) with a compound of Formula (J).

A compound of Formula (AU) where R³ is hydrogen can be prepared bysubjecting a compound of Formula (AS) to Barton-McCombie conditions,such as propanethioyl chloride followed by azobis-isobutyronitrile(AiBN) and tributyltin hydride. A compound of Formula (AV) where R³ ishydrogen can be prepared by removing the t-butyloxycarbonyl (Boc)protecting group from a compound of Formula (AU). Suitable deprotectionreagents include, but are not limited to, trifluoroacetic acid (TFA),hydrochloric acid, and sulfuric acid. A compound of Formula (AW) whereR³ is hydrogen can be prepared by reacting a compound of Formula (AV)with a compound of Formula (J).

Methods for preparing a compound of Formula (AX) are shown in Scheme 10.In Scheme 10, Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵, R⁷ and n can be the sameas Z¹, Z², Z³, Z⁴, R¹, R², R³, R⁵, R⁷ and n as described herein forFormula (I). In Scheme 10, the compounds can be a racemic mixture ofenantiomers or diastereomers. In some embodiments, the compounds can bestereospecific, with carbons ‘a’, ‘b’ and ‘c’ each independently havingeither an (R) or (S) configuration. The compound of Formula (AX) can bea compound of Formula (I), where R⁴ of Formula (I) is

and R⁶ is —NHR⁷. As shown in Scheme 10, a compound of Formula (AX) canbe prepared by reacting a compound of Formula (M) with R⁷—NH₂.

VII. Pharmaceutical Compositions

Some embodiments described herein relate to a pharmaceutical compositionthat can include a therapeutically effective amount of one or morecompounds described herein (e.g., a compound of Formulae (I), or apharmaceutically acceptable salt thereof) and a pharmaceuticallyacceptable carrier, diluent, excipient or combination thereof.Acceptable carriers and diluents for therapeutic use are well known inthe pharmaceutical art, and are described, for example, in Remington'sPharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985),the disclosure of which is incorporated herein by reference in itsentirety. A carrier can facilitate the incorporation of a compound intocells or tissues. For example, without limitation, dimethyl sulfoxide(DMSO) is a commonly utilized carrier that facilitates the uptake ofmany organic compounds into cells or tissues of a subject. A diluent maybe used to increase the bulk of a potent drug whose mass is too smallfor manufacture and/or administration. It may also be a liquid for thedissolution of a drug to be administered by injection, ingestion orinhalation. A common form of diluent in the art is a buffered aqueoussolution such as, without limitation, phosphate buffered saline thatmimics the composition of human blood. Preservatives, stabilizers, dyesand even flavoring agents may be provided in the pharmaceuticalcomposition. For example, sodium benzoate, ascorbic acid and esters ofp-hydroxybenzoic acid may be added as preservatives. In addition,antioxidants and suspending agents may be used.

In some embodiments, the pharmaceutical composition can include a singlediastereomer of a compound of Formula (I)-(VI) (which can include acompound of Formula (Ia), (Ib), (Ic), or (Id)), or a pharmaceuticallyacceptable salt thereof, (for example, a single diastereomer is presentin the pharmaceutical composition at a concentration of greater than 99%compared to the total concentration of the other diastereomers). Inother embodiments, the pharmaceutical composition can include a mixtureof diastereomers of a compound of Formula (I), or a pharmaceuticallyacceptable salt thereof. For example, the pharmaceutical composition caninclude a concentration of one diastereomer of >50%, ≧60%, ≧70%, ≧80%,≧90%, ≧95%, or ≧98%, as compared to the total concentration of the otherdiastereomers. In some embodiments, the pharmaceutical compositionincludes a 1:1 mixture of two diastereomers of a compound of Formula(I), or a pharmaceutically acceptable salt thereof.

A pharmaceutical composition can facilitate administration of one ormore compounds described herein to an organism. Pharmaceuticalcompositions can be obtained by reacting compounds described herein withinorganic or organic acids such as hydrochloric acid, hydrobromic acid,sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid,ethanesulfonic acid, p-toluenesulfonic acid and salicylic acid.Pharmaceutical compositions will generally be tailored to the specificintended route of administration.

The pharmaceutical compositions described herein can be administered toa human patient per se, or in pharmaceutical compositions where they aremixed with other active ingredients, as in combination therapy, orcarriers, diluents, excipients or combinations thereof. Properformulation is dependent upon the route of administration chosen.Techniques for formulation and administration of the compounds describedherein are known to those skilled in the art.

The compounds and compositions disclosed herein may be formulated andused as tablets, capsules, or elixirs for oral administration;suppositories for rectal administration; sterile solutions, suspensionsfor injectable administration; patches for transdermal administration,and sub-dermal deposits and the like. Injectables can be prepared inconventional forms, either as liquid solutions or suspensions, solidforms suitable for solution or suspension in liquid prior to injectionor infusion, or as emulsions. Suitable excipients are, for example,water, saline, dextrose, mannitol, lactose, lecithin, albumin, sodiumglutamate, cysteine hydrochloride, human serum albumin and the like. Inaddition, if desired, the injectable pharmaceutical compositions maycontain minor amounts of nontoxic auxiliary substances, such as wettingagents, pH buffering agents, and the like. If desired, absorptionenhancing preparations (for example, liposomes), may be utilized.

Pharmaceutical formulations for parenteral administration includeaqueous solutions of the compounds in water-soluble form. Additionally,suspensions of the compounds may be prepared as appropriate oilyinjection suspensions. Suitable lipophilic solvents or vehicles includefatty oils such as sesame oil, or other organic oils such as soybean,grapefruit or almond oils, or synthetic fatty acid esters, such as ethyloleate or triglycerides, or liposomes. Aqueous injection suspensions maycontain substances which increase the viscosity of the suspension, suchas sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, thesuspension may also contain suitable stabilizers or agents that increasethe solubility of the compounds to allow for the preparation of highlyconcentrated solutions.

Pharmaceutical preparations for oral use may be obtained by combiningthe compounds with solid excipient, optionally grinding a resultingmixture, and processing the mixture of granules, after adding suitableauxiliaries, if desired, to obtain tablets or dragee cores. Suitableexcipients are, in particular, fillers such as sugars, includinglactose, sucrose, mannitol, or sorbitol; cellulose preparations such as,for example, maize starch, wheat starch, rice starch, potato starch,gelatin, gum tragacanth, methyl cellulose,hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/orpolyvinylpyrrolidone (PVP). If desired, disintegrating agents may beadded, such as the cross-linked polyvinyl pyrrolidone, agar, or alginicacid or a salt thereof such as sodium alginate. Dragee cores areprovided with suitable coatings. For this purpose, concentrated sugarsolutions may be used, which may optionally contain gum arabic, talc,polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/ortitanium dioxide, lacquer solutions, and suitable organic solvents orsolvent mixtures. Dyestuffs or pigments may be added to the tablets ordragee coatings for identification or to characterize differentcombinations of active compound doses. For this purpose, concentratedsugar solutions may be used, which may optionally contain gum arabic,talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/ortitanium dioxide, lacquer solutions, and suitable organic solvents orsolvent mixtures. Dyestuffs or pigments may be added to the tablets ordragee coatings for identification or to characterize differentcombinations of active compound doses. Such formulations can be madeusing methods known in the art.

The pharmaceutical compositions disclosed herein may be manufactured ina manner that is itself known, e.g., by means of conventional mixing,dissolving, granulating, dragee-making, levigating, emulsifying,encapsulating, entrapping or tableting processes. Additionally, theactive ingredients are contained in an amount effective to achieve itsintended purpose. Many of the compounds used in the pharmaceuticalcombinations disclosed herein may be provided as salts withpharmaceutically compatible counterions.

Multiple techniques of administering a compound exist in the artincluding, but not limited to, oral, rectal, topical, aerosol, injectionand parenteral delivery, including intramuscular, subcutaneous,intravenous, intramedullary injections, intrathecal, directintraventricular, intraperitoneal, intranasal and intraocularinjections.

One may also administer the compound in a local rather than systemicmanner, for example, via injection of the compound directly into theinfected area, often in a depot or sustained release formulation.Furthermore, one may administer the compound in a targeted drug deliverysystem, for example, in a liposome coated with a tissue-specificantibody. The liposomes will be targeted to and taken up selectively bythe organ.

The compounds or compositions may, if desired, be presented in a packageor dispenser device which may contain one or more unit dosage formscontaining the active ingredient. The compound or composition describedherein can be packaged alone, or can be packaged with another compoundor another ingredient or additive. The package can contain one or morecontainers filled with one or more of the ingredients of thepharmaceutical compositions. The package may for example comprise metalor plastic foil, such as a blister pack. The package or dispenser devicemay be accompanied by instructions for administration, such asinstructions for administering the compounds or compositions fortreating a neoplastic disease. The package or dispenser may also beaccompanied with a notice associated with the container in formprescribed by a governmental agency regulating the manufacture, use, orsale of pharmaceuticals, which notice is reflective of approval by theagency of the form of the drug for human or veterinary administration.Such notice, for example, may be the labeling approved by the U.S. Foodand Drug Administration for prescription drugs, or the approved productinsert. Compositions that can include a compound described hereinformulated in a compatible pharmaceutical carrier may also be prepared,placed in an appropriate container, and labeled for treatment of anindicated condition.

VIII. Methods of Use

Some embodiments disclosed herein relate to a method of treating and/orameliorating a disease or condition that can include administering to asubject a therapeutically effective amount of one or more compoundsdescribed herein, such as a compound of Formula (I)-(VI) (which caninclude a compound of Formula (Ia), (Ib), (Ic), or (Id)), or apharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition that includes a compound described herein. In someembodiments, the disease or condition can be a neoplastic disease, anage-related disease, a neurological disease, an immunological disease,or an infectious disease. In an embodiment, the neoplastic disease canbe cancer. In some embodiments, the cancer can be B-cell relatedcancers. In some embodiments, the neoplastic disease can be a tumor suchas a solid tumor. In some embodiments, the cancer can be melanoma. Insome embodiments, the cancer can be, breast cancer or pancreatic cancer.In some embodiments, the cancer can be multiple myeloma. In someembodiments, the cancer can be non-Hodgkin's lymphoma. In someembodiment, the cancer can be T cell acute lympho-blastic leukemia. Insome embodiments, the cancer can be mantel cell lymphoma. In someembodiments, the tumor can be glioma. In some embodiments, the specificdiseases described herein (e.g., melanoma, breast cancer, pancreaticcancer, multiple myeloma, non-Hodgkin's lymphoma, T cell acutelympho-blastic leukemia, mantel cell lymphoma or glioma) can beameliorated by a compound of Formula (I)-(VI) (which can include acompound of Formula (Ia), (Ib), (Ic), or (Id)), or a pharmaceuticallyacceptable salt thereof, or a pharmaceutical composition that includes acompound described herein through inhibiting the ubiquitin-proteasomesystem. In some embodiments, the cancer can be a type with low levels oflet-7 microRNA expression, such as lung, colon or breast cancer.

Some embodiments disclosed herein relates to a method for inhibiting theubiquitin-proteasome system in a subject that can include administeringto a subject a therapeutically effective amount of one or more compoundsdescribed herein, such as a compound of Formula (I)-(VI) (which caninclude a compound of Formula (Ia), (Ib), (Ic), or (Id)), or apharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition that includes a compound described herein to inhibit theubiquitin-proteasome system in said subject.

Some embodiments disclosed herein relates to a method for inhibitingCdc34 in a subject that can include administering to a subject atherapeutically effective amount of one or more compounds describedherein, such as a compound of Formula (I)-(VI) (which can include acompound of Formula (Ia), (Ib), (Ic), or (Id)), or a pharmaceuticallyacceptable salt thereof, or a pharmaceutical composition that includes acompound described herein to inhibit Cdc34 in said subject. In oneembodiment, Cdc34 can be hCdc34.

Some embodiments disclosed herein relate to a method for inhibitingcellular proliferation in a subject that can include administering tothe subject a therapeutically effective amount of one or more compoundscompound described herein, such as a compound of Formula (I)-(VI) (whichcan include a compound of Formula (Ia), (Ib), (Ic), or (Id)), or apharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition thereof that includes a compound described herein to inhibitcellular proliferation in said subject.

Some embodiments disclosed herein relate to a method for identifying acandidate therapeutic compound that can include determining theeffective amount of a compound described herein, such as a compound ofFormula (I)-(VI) (which can include a compound of Formula (Ia), (Ib),(Ic), or (Id)), on the extent of ubiquitination of p27^(Kip1) by aSCF^(Skp2) E3 complex, wherein said compound is identified as acandidate therapeutic compound if said compound significantly reducessaid extent of ubiquitination.

Some embodiments disclosed herein relate to a method for determining theeffect of a candidate therapeutic compound include determining theeffective amount of a compound described herein, such as a compound ofFormula (I)-(VI) (which can include a compound of Formula (Ia), (Ib),(Ic), or (Id)), on the extent of ubiquitin chain initiation or ubiquitinchain length, wherein said compound is identified as a candidatetherapeutic compound if said compound significantly reduces said extentof ubiquitin chain initiation or ubiquitin chain length.

Some embodiments disclosed herein relate to a method for determining theeffect of a candidate therapeutic compound that can include determiningthe effective amount of a compound described herein, such as a compoundof Formula (I)-(VI) (which can include a compound of Formula (Ia), (Ib),(Ic), or (Id)), on the extent of cellular proliferation, wherein saidcompound is identified as a candidate therapeutic compound if saidcompound significantly reduces said extent of cellular proliferation.

Some embodiments disclosed herein relate to a method of ameliorating ortreating a neoplastic disease that can include administering to asubject suffering from a neoplastic disease a therapeutically effectiveamount of one or more compounds described herein (e.g., a compound ofFormula (I)-(VI) (which can include a compound of Formula (Ia), (Ib),(Ic), or (Id)), or a pharmaceutically acceptable salt thereof, or apharmaceutical composition that includes a compound described herein).In an embodiment, the neoplastic disease can be cancer. In someembodiments, the neoplastic disease can be a tumor such as a solidtumor. In some embodiments, the cancer can be melanoma, breast cancer orpancreatic cancer. In some embodiments, the cancer can be multiplemyeloma. In some embodiments, the cancer can be non-Hodgkin's lymphoma.In some embodiments, the cancer can be T cell acute lympho-blasticleukemia. In some embodiments, the cancer can be a type with low levelsof let-7 microRNA expression such as lung, colon or breast cancer. Somestudies have suggested that Cdc34 protein levels can be stronglydown-regulated by let-7 overexpression. Reporter assays has demonstrateddirect regulation of the cdc34 3′-untranslated region by let-7. See,Legesse-Miller et al, “let-7 Overexpression leads to an increasedfraction of cells in G2/M, direct down-regulation of Cdc34, andstabilization of Wee1 kinase in primary fibroblasts,” J. Biol. Chem.2009, 284(11):6605-6609.

Some embodiments disclosed herein relate to a method of inhibiting thegrowth of a tumor that can include administering to a subject having atumor a therapeutically effective amount of one or more compoundsdescribed herein (for example, a compound of Formula (I)), or apharmaceutical composition that includes one or more compounds describedherein. In some embodiments, the tumor can be glioma.

In any of the method of treatment described herein by administration ofa compound of Formula (I)-(VI) (which can include a compound of Formula(Ia), (Ib), (Ic), or (Id)) described herein, the method may furthercomprise administering at least one additional therapeutic agent inaddition to the compounds of Formula (I)-(VI) (which can include acompound of Formula (Ia), (Ib), (Ic), or (Id)). In some embodiments, theadditional therapeutic agent is bortezomib (Velcade®). Velcade® is thefirst proteasome inhibitor to reach clinical use as a chemotherapyagent. Bortezomib is used in the treatment of multiple myeloma. In someembodiments, the additional therapeutic agent is ritonavir. Ritonovirhas been shown to inhibit proteasomes as well as free proteases. Somestudies indicate that ritonavir may have inhibitory effects on thegrowth of glioma cells. In some embodiment, the additional therapeuticagent is cisplatin. It has been reported that cisplatin increased ATF5protein expression via preventing its ubiquitin-dependent degradation,which might be associated with its promoting the nucleus-to-cytoplasmtranslocation of E2 ubiquitin-conjugating enzyme Cdc34 and reducing theinteraction between ATF5 and Cdc34. A down-regulation ofproteasome-mediated degradation of ATF5 might contribute tocisplatin-induced apoptosis, providing a new mechanism ofcisplatin-induced apoptosis. See, Wei et al. “Cdc34-mediated degradationof ATF5 is blocked by cisplatin,” J. Biol. Chem. 2008, 283(27):18773-81.

A therapeutically effective amount of a compound disclosed herein canprevent, alleviate or ameliorate symptoms of disease or prolong thesurvival of the subject being treated. This response may occur in atissue, system, animal or human and includes alleviation of the signs orsymptoms of the disease being treated. Determination of atherapeutically effective amount is well within the capability of thoseskilled in the art, in view of the disclosure provided herein. Thetherapeutically effective amount of the compounds disclosed hereinrequired as a dose will depend on the route of administration, the typeof animal, including human, being treated, and the physicalcharacteristics of the specific animal under consideration. The dose canbe tailored to achieve a desired effect, but will depend on such factorsas weight, diet, concurrent medication and other factors which thoseskilled in the medical arts will recognize.

As will be readily apparent to one skilled in the art, the useful invivo dosage to be administered and the particular mode of administrationwill vary depending upon the age, weight, the severity of theaffliction, and mammalian species treated, the particular compoundsemployed, and the specific use for which these compounds are employed.The determination of effective dosage levels, that is the dosage levelsnecessary to achieve the desired result, can be accomplished by oneskilled in the art using routine methods, for example, human clinicaltrials and in vitro studies.

The dosage may range broadly, depending upon the desired effects and thetherapeutic indication. Alternatively dosages may be based andcalculated upon the surface area of the patient, as understood by thoseof skill in the art. Although the exact dosage will be determined on adrug-by-drug basis, in most cases, some generalizations regarding thedosage can be made. The daily dosage regimen for an adult human patientmay be, for example, an oral dose of between 0.01 mg and 3000 mg of eachactive ingredient, preferably between 1 mg and 700 mg, e.g. 5 to 200 mg.The dosage may be a single one or a series of two or more given in thecourse of one or more days, as is needed by the subject. In someembodiments, the compounds will be administered for a period ofcontinuous therapy, for example for a week or more, or for months oryears. In some embodiments, a compound of Formula (I), or apharmaceutically acceptable salt thereof, can be administered lessfrequently compared to the frequency of administration of an agentwithin the standard of care. In some embodiments, a compound of Formula(I), or a pharmaceutically acceptable salt thereof, can be administeredone time per day. For example, a compound of Formula (I), or apharmaceutically acceptable salt thereof, can be administered one timeper day to a subject suffering from a neoplastic disease. In someembodiments, the total time of the treatment regime with a compound ofFormula (I), or a pharmaceutically acceptable salt thereof, can lesscompared to the total time of the treatment regime with the standard ofcare.

In instances where human dosages for compounds have been established forat least some condition, those same dosages may be used, or dosages thatare between about 0.1% and 500%, more preferably between about 25% and250% of the established human dosage. Where no human dosage isestablished, as will be the case for newly-discovered pharmaceuticalcompositions, a suitable human dosage can be inferred from ED₅₀ or ID₅₀values, or other appropriate values derived from in vitro or in vivostudies, as qualified by toxicity studies and efficacy studies inanimals.

In cases of administration of a pharmaceutically acceptable salt,dosages may be calculated as the free base. As will be understood bythose of skill in the art, in certain situations it may be necessary toadminister the compounds disclosed herein in amounts that exceed, oreven far exceed, the above-stated, preferred dosage range in order toeffectively and aggressively treat particularly aggressive diseases orinfections.

Dosage amount and interval may be adjusted individually to provideplasma levels of the active moiety which are sufficient to maintain themodulating effects, or minimal effective concentration (MEC). The MECwill vary for each compound but can be estimated from in vitro data.Dosages necessary to achieve the MEC will depend on individualcharacteristics and route of administration. However, HPLC assays orbioassays can be used to determine plasma concentrations. Dosageintervals can also be determined using MEC value. Compositions should beadministered using a regimen which maintains plasma levels above the MECfor 10-90% of the time, preferably between 30-90% and most preferablybetween 50-90%. In cases of local administration or selective uptake,the effective local concentration of the drug may not be related toplasma concentration.

The exact formulation, route of administration and dosage can be chosenby the individual physician in view of the patient's condition. See forexample, Fingl et al., in The Pharmacological Basis of Therapeutics,1975, the disclosure of which is incorporated herein by reference in itsentirety. It should be noted that the attending physician would know howto and when to terminate, interrupt, or adjust administration due totoxicity or organ dysfunctions. Conversely, the attending physicianwould also know to adjust treatment to higher levels if the clinicalresponse were not adequate (precluding toxicity). The magnitude of anadministrated dose in the management of the disorder of interest willvary with the severity of the condition to be treated and to the routeof administration. The severity of the condition may, for example, beevaluated, in part, by standard prognostic evaluation methods. Further,the dose and perhaps dose frequency, will also vary according to theage, body weight, and response of the individual patient. Determinationof the effective amount of a compound disclosed herein is well withinthe capability of those skilled in the art. A program comparable to thatdiscussed above may be used in veterinary medicine.

Compounds disclosed herein can be evaluated for efficacy and toxicityusing known methods. For example, the toxicology of a particularcompound, or of a subset of the compounds, sharing certain chemicalmoieties, may be established by determining in vitro toxicity towards acell line, such as a mammalian, and preferably human, cell line. Theresults of such studies are often predictive of toxicity in animals,such as mammals, or more specifically, humans. Alternatively, thetoxicity of particular compounds in an animal model, such as mice, rats,rabbits, or monkeys, may be determined using known methods. The efficacyof a particular compound may be established using several recognizedmethods, such as in vitro methods, animal models, or human clinicaltrials. When selecting a model to determine efficacy, the skilledartisan can be guided by the state of the art to choose an appropriatemodel, dose, route of administration and/or regime.

EXAMPLES

Additional embodiments are disclosed in further detail in the followingexamples, which are not in any way intended to limit the scope of theclaims.

Example 1

The following compounds can be prepared according to the reactionconditions shown below.

Example 2

The following compounds can be prepared according to the reactionconditions shown below.

Example 3

The following compounds can be prepared according to the reactionconditions shown below.

Example 4

The following compounds can be prepared according to the reactionconditions shown below.

Example 5

The following compounds can be prepared according to the reactionconditions shown below.

Example 6

The following compounds can be prepared according to the reactionconditions shown below.

Example 7

The following compounds can be prepared according to the reactionconditions shown below.

Example 8

The following compounds can be prepared according to the reactionconditions shown below.

Example 9

The following compounds can be prepared according to the reactionconditions shown below.

Example 10

The following compounds can be prepared according to the reactionconditions shown below.

Example 11

The following compounds can be prepared according to the reactionconditions shown below.

Example 12

The following compounds can be prepared according to the reactionconditions shown below.

Example 13

The following compounds can be prepared according to the reactionconditions shown below.

Example 14

The following compounds can be prepared according to the reactionconditions shown below.

Example 15

The following compounds can be prepared according to the reactionconditions shown below.

Example 16

The following compounds can be prepared according to the reactionconditions shown below.

Example 17 Scheme I

The following Boc-protected compounds were prepared according tosynthetic Scheme I as shown below.

To the solution of DL-p-Bromophenylalanine (I-1) (24.40 g, 100 mmol) in350 mL of anhydrous tetrahydrofuran was added sodium borohydride (15.20g, 400 mmol) in small portions at 0° C. The solution was allowed to stirfor 30 min. Iodine (50.80 g, 200 mmol) was then dissolved in 85 mL ofanhydrous tetrahydrofuran and added dropwise to the original reactionflask slowly in an ice water bath. After stirred for additional 30 min,the resulting mixture was heated to reflux for 30 hrs, cooled to roomtemperature and added dropwise 300 mL of 20% KOH. The resulting solutionwas stirred at 50° C. for 2 hrs, cooled to room temperature, andextracted with dichloromethane (3×300 mL). The combined organic extractswere dried by Na₂SO₄. Removal of the solvent in vacuo afforded a residueI-2 that was used in the next step without further purification.

To the above residue in 200 mL of anhydrous MeOH was added dropwisedi-tert-butyl dicarbonate (33.00 g, 150 mmol) slowly in an ice waterbath. After stirred for additional 30 min, the resulting mixture washeated to 50° C. for 1 h. Removal of the solvent in vacuo afforded whitesolid that was washed with petroleum ether twice to give the desiredproduct I-3 as a white solid (23.00 g, 70%) which was pure enough forthe next reaction. ¹H-NMR (400 MHz, DMSO-d₆): δ 7.46 (d, J=8.4 Hz, 2H),7.16 (d, J=8 Hz, 2H), 6.62 (d, J=8.4 Hz, 1H), 3.56 (brs, 1H), 3.36-3.23(m, 2H), 2.83-2.78 (m, 1H), 2.53 (d, J=11.2 Hz, 1H), 1.30 (s, 9H).

To the solution of I-3 (23.00 g, 70 mmol) and TEMPO (0.47 g, 3 mmol) in750 mL of anhydrous dichloromethane was added trichloroisocyanuric acid(17.44 g, 75 mmol) in small portions at 0° C. After stirred foradditional 20 min, the resulting mixture was filtered and washed withdichloromethane. The filtrate was washed with saturated sodiumthiosulfate (400 mL) and saturated brine (400 mL). The organic extractwas added ethyl (triphenylphosphoranylidene) acetate and refluxed for 2hrs. Removal of the solvent in vacuo afforded a residue that waspurified by flash column chromatography to afford I-5 (15.50 g, 59%) asa white solid. ¹H-NMR (400 MHz, DMSO-d₆): δ 7.48 (d, J=8 Hz, 2H), 7.20(d, J=8 Hz, 3H), 6.88 (dd, J=16, 5.6 Hz, 1H), 5.85 (d, J=15.2 Hz, 1H),4.34 (brs, 1H), 4.14 (q, J=7.2 Hz, 2H), 2.88-2.83 (m, 1H), 2.71-2.64 (m,1H), 1.30 (s, 9H), 1.20 (t, J=7.2 Hz, 3H).

Compound I-5A was prepared in 65% yield following the general proceduredescribed in the synthesis of I-5. ¹H-NMR (400 MHz, DMSO-d₆): δ 7.48 (d,J=8 Hz, 2H), 7.20 (d, J=8.4 Hz, 2H), 7.16 (s, 1H), 6.88 (dd, J=15.6, 5.6Hz, 1H), 5.85 (m, 1H), 4.35 (brs, 1H), 4.14 (q, J=6.8 Hz, 2H), 2.88-2.83(m, 1H), 2.68-2.62 (m, 1H), 1.30 (s, 9H), 1.22 (t, J=7.2 Hz, 3H).

To the solution of I-5 (21.54 g, 57 mmol) and 3,5-dichlorophenylboronicacid (13.00 g, 68 mmol) in 260 mL of N,N-dimethylformamide was addedPd(dppf)Cl₂ (0.64 g) and 2 M Na₂CO₃(75.00 g) under nitrogen. Theresulting mixture was stirred at 95° C. for 6 hrs, cooled to roomtemperature, poured into 600 mL of water, extracted with ethyl acetate(2×400 mL) and washed with saturated brine. Removal of the solvent invacuo afforded a residue that was purified by flash columnchromatography to afford I-6 (15.00 g, 56%) as a white solid. ¹H-NMR(400 MHz, DMSO-d₆): δ 7.71-7.67 (m, 4H), 7.57 (s, 1H), 7.36 (d, J=8.0Hz, 2H), 7.23 (d, J=8.4 Hz, 1H), 6.92 (dd, J=15.6, 5.2 Hz, 1H), 5.88 (d,J=15.6 Hz, 1H), 4.40 (brs, 1H), 4.15 (q, J=6.8 Hz, 2H), 2.94-2.90 (m,1H), 2.77-2.72 (m, 1H), 1.31 (s, 9H), 1.23 (t, J=6.8 Hz, 3H).

I-6A was prepared in 60% yield following the general procedure describedin the synthesis of I-6. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.72-7.67 (m, 4H),7.57 (s, 1H), 7.36 (d, J=8.0 Hz, 2H), 7.23 (d, J=8.8 Hz, 1H), 6.91 (dd,J=15.6, 5.2 Hz, 1H), 5.88 (d, J=15.6 Hz, 1H), 4.40 (brs, 1H), 4.15 (q,J=7.2 Hz, 2H), 2.94-2.90 (m, 1H), 2.77-2.71 (m, 1H), 1.31 (s, 9H), 1.23(t, J=6.8 Hz, 3H).

To the solution of I-6 (9.28 g, 20 mmol) in 60 mL of tetrahydrofuran, 60mL of acetone and 60 mL of water was added 0.1 M OsO₄ in toluene (6 mL)in an ice water bath. The resulting mixture was stirred for additional15 min. NMO (4.68 g, 40 mmol) was added to the reaction, stirred at roomtemperature for overnight, poured into diluted solution of sodiumthiosulfate (300 mL), extracted with ethyl acetate (2×300 mL) and washedwith saturated brine. Removal of the solvent in vacuo afforded a residuethat was purified by flash column chromatography to afford I-7(7.40 g,74%) as a white solid. ¹H-NMR (400 MHz, DMSO-d₆) δ7.70-7.64 (m, 4H),7.56 (s, 1H), 7.30 (d, J=7.6 Hz, 2H), 6.77 (d, J=8.8 Hz, 0.5H), 6.51 (d,J=8.8 Hz, 0.5H), 4.14-4.05 (m, 3H), 3.75-3.67 (m, 2H), 3.12-2.62 (m,2H), 1.28-1.13 (m, 12H).

To the solution of I-6A (4.64 g, 10 mmol) in 40 mL of tetrahydrofuran,40 mL of acetone and 40 mL of water was added (DHQ)₂PHAL (0.78 g, 1mmol) and 0.1 M OsO₄ in toluene (3 mL) in an ice water bath. Theresulting mixture was stirred for additional 15 min. NMO (2.34 g, 20mmol) was added to the reaction, stirred in an ice water bath forovernight, poured into diluted solution of sodium thiosulfate (100 mL),extracted with ethyl acetate (2×200 mL) and washed with saturated brine.Removal of the solvent in vacuo afforded a residue that was purified byflash column chromatography to afford I-7 (3.00 g, 60%) as a whitesolid. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.69-7.64 (m, 4H), 7.55 (s, 1H), 7.29(d, J=5.2 Hz, 2H), 6.74 (d, J=6.0 Hz, 1H), 5.00 (brs, 1H), 4.95 (brs,1H), 4.14-4.10 (m, 3H), 3.76-3.69 (m, 2H), 3.11-3.09 (m, 1H), 2.64-2.60(m, 1H), 1.28-1.13 (m, 12H).

General Procedure for the Synthesis of Active Esters:

To the solution of acid (30 mmol) and N-hydroxysuccinimide (3.68 g, 32mmol) in 120 mL of dichloromethane was slowly added DCC (6.60 g, 32mmol) in small portions in an ice water bath. The reaction was stirredfor additional 2 hrs, filtered with a pad of silica-gel, washed withdichloromethane. Removal of the solvent in vacuo afforded the desiredproduct which was used in the next step without further purification.

To the solution of I-7 (1.00 g, 2 mmol) in 8 mL of dichloromethane and 8mL of ethanol was added 6 M HCl in dioxane (6 mL). The reaction wasstirred at 30° C. for 2 hrs. Removal of the solvent in vacuo afforded aresidue that was dissolved in 10 mL of N,N-dimethylformamide in an icewater bath. 14% Na₂CO₃ (3.03 g, 4 mmol) and active ester (3 mmol) wasadded to the reaction, stirred at room temperature for 3 hrs, pouredinto diluted solution of ammonium chloride (100 mL), extracted withethyl acetate (2×100 mL) and washed with saturated brine. Removal of thesolvent in vacuo afforded a residue that was purified by flash columnchromatography to afford I-8.

A1 was prepared in 64% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.83 (d, J=9.2 Hz,0.5H), 7.72-7.65 (m, 4H), 7.56 (d, J=1.6 Hz, 1H), 7.45 (d, J=8.8 Hz,0.5H), 7.30 (d, J=7.6 Hz, 2H), 4.16-4.04 (m, 4H), 3.75-3.64 (m, 3H),3.24 (s, 1.5H), 3.20 (s, 1.5H), 3.12-2.79 (m, 2H), 1.22-1.14 (m, 3H).

A2 was prepared in 60% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.83 (d, J=9.2 Hz,1H), 7.72-7.65 (m, 4H), 7.56 (s, 1H), 7.31 (d, J=8.0 Hz, 2H), 4.16-4.10(m, 4H), 3.79-3.60 (m, 3H), 3.20 (s, 3H), 3.12-3.07 (m, 1H), 2.82-2.76(m, 1H), 1.23 (t, J=7.2 Hz, 3H).

B1 was prepared in 70% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.72-7.65 (m, 4H),7.56 (s, 1H), 7.37-7.28 (m, 3H), 4.17-4.03 (m, 4H), 3.78-3.70 (m, 3H),3.39-3.31 (m, 2H), 2.90-2.79 (m, 2H), 1.21-1.08 (m, 6H).

B2 was prepared in 55% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 8.09-7.56 (m, 6H),7.29-7.23 (m, 4H), 6.93-6.84 (m, 3H), 4.41-4.36 (m, 2H), 4.18-4.06 (m,4H), 3.78-3.77 (m, 1H), 2.89-2.73 (m, 2H), 1.22-1.15 (m, 3H).

B3 was prepared in 50% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 8.19 (d, J=6.0 Hz,0.4H), 8.02 (d, J=5.6 Hz, 0.6H), 7.72-7.56 (m, 5H), 7.30 (d, J=5.2 Hz,2H), 5.35-5.07 (m, 2H), 4.08-3.95 (m, 6H), 3.25 (brs, 1H), 3.10-2.73 (m,2H), 1.21-1.15 (m, 3H).

B4 was prepared in 58% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.73-7.64 (m, 5H),7.55 (S, 1H), 7.29-7.27 (m, 2H), 4.15-4.06 (m, 5H), 3.78-3.69 (m, 3H),3.15-2.78 (m, 2H), 1.40-1.16 (m, 7H).

B5 was prepared in 66% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 8.09-7.56 (m, 5H),7.36-7.30 (m, 3H), 4.17-3.70 (m, 6H), 3.28-3.13 (m, 2H), 2.89-2.73 (m,2H), 1.91-1.14 (m, 3H), 0.85-0.82 (m, 13H).

B6 was prepared in 72% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 8.01 (d, J=6.0 Hz,0.5H), 7.69-7.61 (m, 4.5H), 7.56 (s, 1H), 7.30 (m, 2H), 6.81-6.74 (m,1H), 4.23-3.67 (m, 6H), 3.20-3.14 (m, 1H), 2.82-2.68 (m, 1H), 1.21-0.89(m, 15H).

B7 was prepared in 46% yield following the general procedure describedin the synthesis of I-8. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.80-7.62 (m, 5H),7.56 (s, 1H), 7.30 (m, J=5.2 Hz, 2H), 6.84 (s, 1H), 5.07 (brs, 1H), 4.90(brs, 1H), 4.17-3.69 (m, 4H), 3.08-3.05 (m, 1H), 2.83-2.68 (m, 1H), 1.36(s, 9H), 1.21-1.18 (t, J=4.8 Hz, 3H).

General procedure for the synthesis of I-9: To the solution of I-8 (1mmol) in 10 mL of tetrahydrofuran and 0.2 mL of water was added lithiumhydroxide monohydrate (0.21 g, 5 mmol). The reaction was stirred at 35°C. for 1 h, poured into diluted hydrochloric acid (60 mL), extractedwith ethyl acetate (2×60 mL) and washed with saturated brine. Removal ofthe solvent in vacuo afforded a residue that was purified by flashcolumn chromatography to afford the desired product I-9.

A3 was prepared in 40% yield following the general procedure describedin the synthesis of I-9. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.80 (d, J=9.2 Hz,1H), 7.72-7.65 (m, 4H), 7.56 (s, 1H), 7.31 (d, J=7.6 Hz, 1H), 4.16-4.05(m, 2H), 3.78-3.61 (m, 3H), 3.22-3.20 (ss, 3H), 3.11-3.08 (m, 1H),2.82-2.76 (m, 1H).

A4 was prepared in 43% yield following the general procedure describedin the synthesis of I-9. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.80 (d, J=8.8 Hz,1H), 7.72-7.65 (m, 4H), 7.56 (s, 1H), 7.31 (d, J=8.0 Hz, 1H), 5.04 (brs,1H), 4.85 (brs, 1H), 4.17-4.09 (m, 2H), 3.75-3.65 (m, 3H), 3.20 (s, 3H),3.11-3.07 (m, 1H), 2.82-2.76 (m, 1H).

B8 was prepared in 50% yield following the general procedure describedin the synthesis of I-9. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.82-7.61 (m, 4H),7.54 (s, 1H), 7.45-7.30 (m, 3H), 3.92-3.55 (m, 4H), 3.39-3.14 (m, 3H),2.94-2.82 (m, 2H), 1.09-0.99 (m, 3H).

B9 was prepared in 48% yield following the general procedure describedin the synthesis of I-9. ¹H-NMR (400 MHz, DMSO-d₆) δ 8.09-7.56 (m, 6H),7.30-7.23 (m, 4H), 6.92-6.85 (m, 3H), 4.40-4.38 (m, 2H), 4.20-4.11 (m,2H), 3.82-3.80 (m, 1H), 3.09-2.79 (m, 2H).

B10 was prepared in 60% yield following the general procedure describedin the synthesis of I-9. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.69-7.65 (m, 5H),7.56 (s, 1H), 7.29 (brs, 2H), 4.11-4.04 (m, 3H), 3.78-3.70 (m, 3H),3.10-2.78 (m, 2H), 1.98-1.17 (m, 4H).

General Procedure for the Synthesis of Amines from Boc-Amide:

To the solution of amide (250 mg) in 10 mL of ethyl acetate was added5.8 M HCl in ethyl acetate (5 mL). The reaction was stirred at 25° C.for 2 hrs, poured into saturated sodium bicarbonate (60 mL), extractedwith ethyl acetate (2×50 mL) and dried over anhydrous sodium sulfate.Removal of the solvent in vacuo afforded a residue that was purified byflash column chromatography to afford the desired product.

B11 was prepared in 43% yield following the general procedure describedabove. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.71-7.56 (m, 6H), 7.36-7.31 (m, 2H),4.19-4.12 (m, 4H), 3.73-3.65 (m, 1H), 3.39-3.09 (m, 3H), 2.75 (brs, 1H),1.21 (brs, 3H).

B12 was prepared in 55% yield following the general procedure describedabove. ¹H-NMR (400 MHz, DMSO-d₆) δ 7.93-7.64 (m, 5H), 7.55 (s, 1H), 7.31(d, J=4.4 Hz, 2H), 4.17-4.03 (m, 4H), 3.73-3.70 (m, 1H), 3.45-3.08 (m,2H), 2.81-2.75 (m, 1H), 1.21 (brs, 3H), 0.95 (brs, 3H).

General Procedure for the Synthesis of Esters I-10:

To the solution of I-9 (0.9 mmol) in 5 mL of alcohol was added 5.8 M HClin ethyl acetate (5 mL). The reaction was stirred at 25° C. for 3 hrs,poured into saturated sodium bicarbonate (60 mL), extracted with ethylacetate (2×50 mL) and dried over anhydrous sodium sulfate. Removal ofthe solvent in vacuo afforded a residue that was purified by flashcolumn chromatography to afford the desired product I-10.

B13 was prepared in 60% yield following the general procedure describedin the synthesis of I-10. ¹H-NMR (400 MHz, DMSO-d₆): δ7.80 (d, J=6.0 Hz,1H), 7.71-7.65 (m, 4H), 7.55 (s, 1H), 7.30 (d, J=5.2 Hz, 2H), 5.12 (d,J=5.2 Hz, 2H), 4.18-4.14 (m, 2H), 3.80-3.61 (m, 6H), 3.19 (s, 3H),3.11-3.08 (m, 1H), 2.81-2.77 (m, 1H).

B14 was prepared in 57% yield following the general procedure describedin the synthesis of I-10. ¹H-NMR (400 MHz, DMSO-d₆): δ7.70 (d, J=0.8 Hz,2H), 7.65 (d, J=5.2 Hz, 2H), 7.56 (s, 1H), 7.46 (d, J=6.0 Hz, 1H),7.38-7.22 (m, 7H), 5.15-5.05 (m, 3H), 4.17 (brs, 2H), 3.78-3.63 (m, 3H),3.21 (s, 3H), 2.93-2.90 (m, 1H), 2.81-2.78 (m, 1H).

Example 18 Assays Proliferation Assays

Proliferation assays were conducted to assess the effects of thecompounds described herein on the growth rate of neoplastic cells. Foreach of the proliferation assays described herein, SK-MEL-28, MDA-MB-468or Panc-1 cells were grown in RPMI supplemented with 10% FBS and platedonto sterile 96-well tissue culture plates at 20,000 cells per well andallowed to attach overnight. Compounds B1 through B14 were added at 20μM and 10 μM respectively in the SK-MEL-28 proliferation assay (FIG. 1).Compounds B3, B5, B7, B11 and B12 were added at 20 μM and 10 μMrespectively for the MDA-MB-468 (FIG. 2A) and Panc-1 proliferationassays (FIG. 2B). Plates were incubated for 72 h (37° C., 5% CO₂).Percent viability compared to DMSO control was determined using an MTSassay (CellTiter 96 Aqueous, Promega) according to the manufacturer'sstandard protocol. The standard used in the SK-MEL-28 assay is MG132, aproteasome inhibitor, at 10 uM concentration. It was used as a positivecontrol for cell killing. All the assays are referenced to DMSO as thecontrol at 100% proliferation.

In particular, Compound B12 demonstrated excellent inhibitory effect at20 μM concentration, which is about 20 fold more effective than thecontrol (FIG. 1). Compounds B3, B5, B7 and B11 also showed goodinhibitory effect.

Compound B3 demonstrated good inhibitory effect in both the MDA-MB-468assay and the Panc-1 assay at both concentrations, showing 5.5% and 6.0%growth rate in the MDA-MB-468 assay and 5.2% and 4.8% growth rate in thePanc-1 assay at 20 μM and 10 μM respectively. Compound B5 and CompoundB12 showed 33.5% and 11.0% growth rate respectively in the MDA-MD-468assay when used at 20 μM. Compound B12 showed 24.4% growth rate in thePanc-1 assay when used at 20 μM.

TNF-α Induced NFkB Reporter Assays:

Generation of Stable MDA-MB-468-NFkB-Luc2p Reporter Cell Line:

The effects of the compounds described herein on the activation ofTNF-mediated pathways were assessed using an MDA-MB-468NFkB-Luc2preporter cell line (FIG. 3). To construct this cell line, MDA-MB-468cells were transfected with pGL4.32[luc2P/NF-kB-RE/Hygro] vector(Promega) using TransFast transfection reagent (Promega) according tomanufacturer's standard protocol. Briefly, cells were grown in RPMIsupplemented with 10% FBS and plated onto sterile 6-well plates (Nunc).When cells became ˜75% confluent, media in each well was removed andreplaced with 1 mL RPMI (no FBS) containing pGL4.41[luc2P/NFkB/Hygro]vector and TransFast for 1 h at 37° C. 2 mL of RPMI with FBS is thenadded to the well and after 24 h, 500 ug/ml of hygromycin was added forselection to generate a stable reporter cell line.

Assay Results

Stable MDA-MB-468-NFkB-Luc2p cells were plated onto optical-bottom,white wall 96-well plates at 10,000 cells per well and allowed to attachovernight. Compounds B1 through B14 were added at 20 μM and 5 μMrespectively for 1 h prior to treatment with 10 ng/ml of recombinanthuman TNF-α for 6 h. Luciferase signal was determined using theLuciferase Assay System (Promega) and a luminometer. The results areprovided in FIG. 3. In particular, Compound B12 demonstrated excellentinhibitory effect at 20 μM concentration. Compounds B1, B4, and B5 alsoshowed good results.

Furthermore, although the foregoing has been described in some detail byway of illustrations and examples for purposes of clarity andunderstanding, it will be understood by those of skill in the art thatnumerous and various modifications can be made without departing fromthe spirit of the present disclosure. Therefore, it should be clearlyunderstood that the forms disclosed herein are illustrative only and arenot intended to limit the scope of the present disclosure, but rather toalso cover all modification and alternatives coming with the true scopeand spirit of the invention.

What is claimed is:
 1. A compound of Formula (I) or a pharmaceuticallyacceptable salt thereof:

wherein: n is selected from the group consisting of 0, 1, 2, 3, 4, and5; each R¹ is independently selected from the group consisting of halo,cyano, and azido; Z¹, Z², Z³, and Z⁴ are each independently —CH— or —N—;R² is selected from the group consisting of an optionally substituted(C₁₋₆ alkoxy)C₁₋₆ alkyl, an optionally substituted C₆₋₁₀ aryl, anoptionally substituted C₅₋₁₀ heteroaryl, an optionally substituted(aryloxy)C₁₋₆ alkyl, an optionally substituted C₃₋₇ heterocyclyl, anoptionally substituted C₃₋₇ cycloalkyl, an optionally substitutedhaloalkyl, and an optionally substituted aminoalkyl; R³ is hydrogen or—OH; R⁴ is selected from the group consisting of an optionallysubstituted C₃₋₆ cycloalkyl, an optionally substituted C₃₋₆heterocyclyl, an optionally substituted C₆₋₁₀aryl, an optionallysubstituted C₅₋₁₀ heteroaryl,

or R³ and R⁴ together form an optionally substituted C₃₋₆ heterocyclicring; R⁵ is hydrogen or —OH; R⁶ is selected from the group consisting of—OH, —NHR⁷, an optionally substituted C₁₋₆ alkyl, an optionallysubstituted C₁₋₆ alkoxy, an optionally substituted C₆₋₁₀ aryl, anoptionally substituted C₅₋₁₀ heteroaryl, an optionally substitutedaryloxy, and an optionally substituted arylalkoxy; R⁷ is hydrogen or anoptionally substituted C₁₋₆ alkyl; t is selected from the groupconsisting of 0, 1, 2, 3, 4, and 5; provided that if n is 2, both R¹ arechloro, R² is methoxymethyl, R³ is —OH, R⁴ is —CH(OH)CH₂OH or

 R⁵ is —OH, and R⁶ is —OH, —NHR⁷, or methoxy; then at least one of Z¹,Z², Z³, and Z⁴ is —N—; provided that if n is 2, both R¹ are chloro, R²is methoxymethyl or phenyl, R³ is hydrogen, R⁴ is

 R⁵ is hydrogen, and R⁶ is —OH; then at least one of Z¹, Z², Z³, and Z⁴is —N—; provided that if n is 2, both R¹ are chloro, R² ismethoxymethyl, and R³ and R⁴ together form an optionally substitutedheterocyclic ring, then at least one of Z¹, Z², Z³, and Z⁴ is —N—;provided that if n is 2, both R¹ are chloro, R² is methoxymethyl, R³ is—OH, R⁴ is

 R⁵ is —OH, then R⁶ cannot be —OH, —NH(CH₂)₃OCH₃, or —NH(CH₂)₃N(CH₃)₂;provided that if n is 2, both R¹ are chloro, R² is methoxymethyl, R³ is—OH, R⁴ is

 R⁵ is —OH, R⁶ is methoxy, then the compound cannot be


2. The compound of claim 1, wherein n is selected from the groupconsisting of 1, 2, 3, 4, and
 5. 3. The compound of claim 2, wherein nis
 2. 4. The compound of claim 1, wherein each R¹ is halo.
 5. Thecompound of claim 4, wherein each R¹ is chloro.
 6. The compound of claim1, wherein Z¹, Z², Z³ and Z⁴ are each —CH—.
 7. The compound of claim 1,wherein R² is (C₁₋₆ alkoxy)C₁₋₆ alkyl.
 8. The compound of claim 7,wherein R² is methoxymethyl or ethoxymethyl.
 9. The compound of claim 1,wherein R² is (aryloxy)C₁₋₆ alkyl.
 10. The compound of claim 9, whereinR² is phenoxymethyl.
 11. The compound of claim 1, wherein R² is C₃₋₇heterocyclyl.
 12. The compound of claim 11, wherein R² is selected fromoptionally substituted tetrahydrofuranyl or optionally substitutedpyrrolidinyl.
 13. The compound of claim 12, wherein the nitrogen atom inpyrrolidinyl is protected with a t-butyloxycarbonyl (Boc) protectinggroup.
 14. The compound of claim 1, wherein R² is C₃₋₇ cycloalkyl. 15.The compound of claim 14, wherein R² is cyclopentyl.
 16. The compound ofclaim 1, wherein R² is haloalkyl.
 17. The compound of claim 16, whereinR² is selected from the group consisting of —CH₂Cl, —CH₂Br, —CH₂CH₂Cl,—CH₂CH₂Br, —CH(Cl)CH₃ and —CH(Br)CH₃.
 18. The compound of claim 1,wherein R² is optionally substituted aminoalkyl.
 19. The compound ofclaim 18, wherein R² is selected from the group consisting of —CH₂NH₂,—CH₂NH(Boc), —CH(NH₂)CH₃, and —CH(Boc-NH)CH₃.
 20. The compound of claim1, wherein R³ is —OH.
 21. The compound of claim 1, wherein R⁴ is


22. The compound of claim 21, wherein R⁵ is —OH.
 23. The compound ofclaim 1, wherein R⁶ is selected from the group consisting of —OH, —NHR⁷,an optionally substituted C₁₋₆ alkoxy, and an optionally substitutedarylalkoxy.
 24. The compound of claim 23, wherein R⁶ is selected fromthe group consisting of —OH, —OCH₂Ph and —OCH₂CH₃.
 25. The compound ofclaim 23, wherein R⁶ is —OCH₂CH₃.
 26. The compound of claim 23, whereinR⁶ is a substituted C₁₋₆ alkyl.
 27. The compound of claim 26, whereinthe C₁₋₆ alkyl is substituted with one or more groups selected from thegroup consisting of halogen, —OH, —COOH, —NR⁸R⁹, C₁₋₆ alkoxy, and C₅₋₁₀heteroaryl; wherein R⁸ and R⁹ are each independently hydrogen or C₁₋₆alkyl.
 28. The compound of claim 23, wherein R⁶ is —NHR⁷.
 29. Thecompound of claim 28, wherein R⁷ is an optionally substituted C₁₋₆alkyl.
 30. The compound of claim 1, wherein the compound of Formula (I)is selected from the group consisting of Formula (II), Formula (III),Formula (IV), Formula (V) and Formula (VI), or a pharmaceuticallyacceptable salt thereof:

wherein R⁴ is selected from


31. The compound of claim 1, wherein the compound of Formula (I) isselected from the group consisting of:

a pharmaceutically acceptable salt thereof.
 32. The compound of claim30, wherein the compound is enriched with respect to the shownstereochemistry in an amount that is >50% as compared to the amount ofother stereoisomer impurities.
 33. The compound of claim 30, wherein thecompound is enriched with respect to the shown stereochemistry in anamount that is >60% as compared to the amount of other stereoisomerimpurities.
 34. The compound of claim 30, wherein the compound isenriched with respect to the shown stereochemistry in an amount thatis >70% as compared to the amount of other stereoisomer impurities. 35.The compound of claim 30, wherein the compound is enriched with respectto the shown stereochemistry in an amount that is >80% as compared tothe amount of other stereoisomer impurities.
 36. The compound of claim30, wherein the compound is enriched with respect to the shownstereochemistry in an amount that is >90% as compared to the amount ofother stereoisomer impurities.
 37. The compound of claim 30, wherein thecompound is enriched with respect to the shown stereochemistry in anamount that is >95% as compared to the amount of other stereoisomerimpurities.
 38. The compound of claim 30, wherein the compound isenriched with respect to the shown stereochemistry in an amount thatis >98% as compared to the amount of other stereoisomer impurities. 39.A pharmaceutical composition comprising a therapeutically effectiveamount of a compound of claim 1, or a pharmaceutically acceptable saltthereof, and a pharmaceutically acceptable carrier, diluent, excipientor combination thereof.
 40. The pharmaceutical composition of claim 39,wherein the compound is a mixture of two or more diastereomers.
 41. Amethod for inhibiting the ubiquitin-proteasome system in a subjectcomprising administering to the subject a compound of claim 1, or apharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition thereof, in a therapeutically effective amount sufficient toinhibit the ubiquitin-proteasome system in said subject.
 42. A methodfor inhibiting Cdc34 in a subject comprising administering to thesubject a compound of claim 1, or a pharmaceutically acceptable saltthereof, or a pharmaceutical composition thereof, in a therapeuticallyeffective amount sufficient to inhibit Cdc34 in said subject.
 43. Themethod of claim 42, wherein the Cdc34 is hCdc34.
 44. A method forinhibiting cellular proliferation in a subject comprising administeringto the subject a compound of claim 1, or a pharmaceutically acceptablesalt thereof, or a pharmaceutical composition thereof, in atherapeutically effective amount sufficient to inhibit cellularproliferation in said subject.
 45. A method for ameliorating a conditionselected from the group consisting of a neoplastic disease, aneurological disease, an immunological disease, and an infectiousdisease, comprising administering to a subject suffering from thecondition a therapeutically effective amount of a compound of claim 1,or a pharmaceutically acceptable salt thereof, or a pharmaceuticalcomposition thereof.
 46. The method of claim 45, wherein the neoplasticdisease is cancer.
 47. The method of claim 46, wherein the cancer isselected from the group consisting of melanoma, breast cancer,pancreatic cancer, multiple myeloma, mantle cell lymphoma, glioma,cancers with low levels of the let-7 microRNA, lung cancer, coloncancer, non-hodgkin's lymphoma, and T cell acute lympho-blasticleukemia.
 48. A method for identifying a candidate therapeutic compoundcomprising determining the effective amount of a compound of claim 1 onthe extent of ubiquitination of p27^(Kip1) by a SCF^(Skp2) E3 complex,wherein said compound is identified as a candidate therapeutic compoundif said compound significantly reduces said extent of ubiquitination.49. A method for determining the effect of a candidate therapeuticcompound comprising determining the effective amount of a compound ofclaim 1 on the extent of ubiquitin chain initiation or ubiquitin chainlength, wherein said compound is identified as a candidate therapeuticcompound if said compound significantly reduces said extent of ubiquitinchain initiation or ubiquitin chain length.
 50. A method for determiningthe effect of a candidate therapeutic compound comprising determiningthe effective amount of a compound of claim 1 on the extent of cellularproliferation, wherein said compound is identified as a candidatetherapeutic compound if said compound significantly reduces said extentof cellular proliferation.